Masoud Rezvani Jalal and Saba Fathollahi
In this paper, the optical force between two circular plasmonic wires of submicrometer diameter (0.3 μm) with nanometer surface-to-surface distances (3–30 nm) interacting with radiation of a complex point source (𝜆≈0.2λ≈0.2 μm) is numerically studied. Calculations (which are based on the Müller integral equations and the Maxwell stress tensor) show that an attractive optical force with a number of distinct peaks is created in distances 3–10 nm. However, for plasmonic–dielectric and plasmonic–reflector double-wires, the optical force has no such peaks. Comparisons reveal that the peaks are originated from the excitation of coupled surface plasmon polaritons in the gap region between the plasmonic wires.
DOI
Friday, June 30, 2017
In situ self-assembly and photopolymerization for hetero-phase synthesis and patterning of conducting materials using soft oxometalates in thermo-optical tweezers
Subhrokoli Ghosh, Santu Das, Shuvojit Paul, Preethi Thomas, Basudev Roy, Partha Mitra, Soumyajit Roy and Ayan Banerjee
We demonstrate a novel method of simultaneous photoassisted hetero-phase synthesis, doping, and micro-scale patterning of conductive materials. The patterning is performed by controlled self-assembly mediated by a micro-bubble induced in an optical tweezers configuration. The high temperature generated due to the light field of the tweezers also drives diverse chemical reactions that lead to the in situ formation of conducting metal-oxides and polymers due to a charge transfer mechanism with soft oxometalates (SOMs). We synthesize two conducting polymers – polypyrrole and polyaniline – doped by the metal oxides Mo–O2 and Mo–O3, from dispersions of the respective organic compounds with the SOMs, and form permanent patterns out of them by continuous self-assembly arising from manipulation of the micro-bubble using Marangoni flows generated by the tweezers. The electrically conducting patterns of width varying between around 4–50 μm, are written in the form of simple Hall-bar geometries, and a four-probe measurement technique yields conductivities on the order of ∼450–600 Siemens cm−1 – which is much higher than that reported for both polypyrrole and polyaniline in earlier work. This technique can easily be used in patterning complicated electrical circuits in mesoscopic length scales, and can also be extended to solution processed electronic device development by green chemical routes.
DOI
We demonstrate a novel method of simultaneous photoassisted hetero-phase synthesis, doping, and micro-scale patterning of conductive materials. The patterning is performed by controlled self-assembly mediated by a micro-bubble induced in an optical tweezers configuration. The high temperature generated due to the light field of the tweezers also drives diverse chemical reactions that lead to the in situ formation of conducting metal-oxides and polymers due to a charge transfer mechanism with soft oxometalates (SOMs). We synthesize two conducting polymers – polypyrrole and polyaniline – doped by the metal oxides Mo–O2 and Mo–O3, from dispersions of the respective organic compounds with the SOMs, and form permanent patterns out of them by continuous self-assembly arising from manipulation of the micro-bubble using Marangoni flows generated by the tweezers. The electrically conducting patterns of width varying between around 4–50 μm, are written in the form of simple Hall-bar geometries, and a four-probe measurement technique yields conductivities on the order of ∼450–600 Siemens cm−1 – which is much higher than that reported for both polypyrrole and polyaniline in earlier work. This technique can easily be used in patterning complicated electrical circuits in mesoscopic length scales, and can also be extended to solution processed electronic device development by green chemical routes.
DOI
Myosin Va molecular motors manoeuvre liposome cargo through suspended actin filament intersections in vitro
Andrew T. Lombardo, Shane R. Nelson, M. Yusuf Ali, Guy G. Kennedy, Kathleen M. Trybus, Sam Walcott & David M. Warshaw
Intracellular cargo transport relies on myosin Va molecular motor ensembles to travel along the cell’s three-dimensional (3D) highway of actin filaments. At actin filament intersections, the intersecting filament is a structural barrier to and an alternate track for directed cargo transport. Here we use 3D super-resolution fluorescence imaging to determine the directional outcome (that is, continues straight, turns or terminates) for an ∼10 motor ensemble transporting a 350 nm lipid-bound cargo that encounters a suspended 3D actin filament intersection in vitro. Motor–cargo complexes that interact with the intersecting filament go straight through the intersection 62% of the time, nearly twice that for turning. To explain this, we develop an in silico model, supported by optical trapping data, suggesting that the motors’ diffusive movements on the vesicle surface and the extent of their engagement with the two intersecting actin tracks biases the motor–cargo complex on average to go straight through the intersection.
DOI
Intracellular cargo transport relies on myosin Va molecular motor ensembles to travel along the cell’s three-dimensional (3D) highway of actin filaments. At actin filament intersections, the intersecting filament is a structural barrier to and an alternate track for directed cargo transport. Here we use 3D super-resolution fluorescence imaging to determine the directional outcome (that is, continues straight, turns or terminates) for an ∼10 motor ensemble transporting a 350 nm lipid-bound cargo that encounters a suspended 3D actin filament intersection in vitro. Motor–cargo complexes that interact with the intersecting filament go straight through the intersection 62% of the time, nearly twice that for turning. To explain this, we develop an in silico model, supported by optical trapping data, suggesting that the motors’ diffusive movements on the vesicle surface and the extent of their engagement with the two intersecting actin tracks biases the motor–cargo complex on average to go straight through the intersection.
DOI
Improvement of Sensing and Trapping Efficiency of Double Nanohole Apertures via Enhancing the Wedge Plasmon Polariton Modes with Tapered Cusps
Mostafa Ghorbanzadeh, Steven Jones, Mohammad Kazem Moravvej-Farshi, and Reuven Gordon
In the past few years, double nanohole (DNH) apertures in a gold film have been used extensively to trap and sense biological and artificial dielectric nanoparticles. Using numerical simulations we show that the conical shape of a DNH, milled by a focused ion beam into a thin gold layer, which is an inherent property of the fabrication process, plays a critical role in the sensitivity of the DNHs, and is beneficial to the optical sensing and trapping applications. The slope of the metallic wedges in an appropriately designed DNH leads to 2D nanofocusing of gap surface plasmons (GSPs) and couples them to the wedge plasmon polaritons (WPPs), creating “hot spots” required for trapping. The transmission variations due to the trapping polystyrene nanoparticles of radii 11 ± 1 nm by particularly designed DNHs, measured at the wavelength near the corresponding wedge mode resonance, are shown to be in good agreements with numerical results using conically modeled DNHs. This observation highlights the extreme sensitivity of aperture assisted trapping, specifically with regard to the DNH structure. These findings open up new routes toward the design and optimization of efficient aperture structures for trapping and sensing applications.
DOI
In the past few years, double nanohole (DNH) apertures in a gold film have been used extensively to trap and sense biological and artificial dielectric nanoparticles. Using numerical simulations we show that the conical shape of a DNH, milled by a focused ion beam into a thin gold layer, which is an inherent property of the fabrication process, plays a critical role in the sensitivity of the DNHs, and is beneficial to the optical sensing and trapping applications. The slope of the metallic wedges in an appropriately designed DNH leads to 2D nanofocusing of gap surface plasmons (GSPs) and couples them to the wedge plasmon polaritons (WPPs), creating “hot spots” required for trapping. The transmission variations due to the trapping polystyrene nanoparticles of radii 11 ± 1 nm by particularly designed DNHs, measured at the wavelength near the corresponding wedge mode resonance, are shown to be in good agreements with numerical results using conically modeled DNHs. This observation highlights the extreme sensitivity of aperture assisted trapping, specifically with regard to the DNH structure. These findings open up new routes toward the design and optimization of efficient aperture structures for trapping and sensing applications.
DOI
Low-loss nanowire and nanotube plasmonic waveguide with deep subwavelength light confinement and enhanced optical trapping forces
Xiaogang Chen, Qijing Lu, Xiang Wu, Hongqin Yang and Shusen Xie
With the rapid development of the micro/nano fabrication technology, the semiconductor nanowires and nanotubes with size and dimensions controllable realize wide applications in nanophotonics. In this talk, we propose two kinds of hybrid plasmonics waveguides, one is consisting of nanowires, another is consisting of nanotubes. By employing the simulating with different geometric parameters, the basic waveguiding properties, including the effective mode area, the propagation length, the mode character and the optical trapping forces can be achieved. Compared with previous plasmonic waveguide with plane metal substrate, current plasmonics waveguides with ease of fabrication have the advantage of long propagation length and effectively optical trapping of nanoparticles with deep subwavelength light confinement, which may be very useful for nanophotonic integrated circuits, nanolasers and biosensing.
DOI
With the rapid development of the micro/nano fabrication technology, the semiconductor nanowires and nanotubes with size and dimensions controllable realize wide applications in nanophotonics. In this talk, we propose two kinds of hybrid plasmonics waveguides, one is consisting of nanowires, another is consisting of nanotubes. By employing the simulating with different geometric parameters, the basic waveguiding properties, including the effective mode area, the propagation length, the mode character and the optical trapping forces can be achieved. Compared with previous plasmonic waveguide with plane metal substrate, current plasmonics waveguides with ease of fabrication have the advantage of long propagation length and effectively optical trapping of nanoparticles with deep subwavelength light confinement, which may be very useful for nanophotonic integrated circuits, nanolasers and biosensing.
DOI
Plasmonic Chiral Nanostructures: Chiroptical Effects and Applications
Yang Luo, Cheng Chi, Meiling Jiang, Ruipeng Li, Shuai Zu, Yu Li, Zheyu Fang
The plasmonic chiroptical effect has been used to manipulate chiral states of light, where the strong field enhancement and light localization in metallic nanostructures can amplify the chiroptical response. Moreover, in metamaterials, the chiroptical effect leads to circular dichroism (CD), circular birefringence (CB), and asymmetric transmission. Potential applications enabled by chiral plasmonics have been realized in various areas of nanoscience and nanotechnology. In this review, both basic theories and state-of-the-art studies on plasmonic chiroptical effects are summarized. Molecular chiroptical effects are drastically enhanced by metallic nanostructures that can generate a “superchiral” field, which arises from the strong electromagnetic interactions. Both intrinsic and extrinsic plasmonic chiral metamaterials formed by the periodic arrangement of metallic nanostructured units show high levels of CB, CD, and asymmetric transmission. Consequent applications including photo detection, molecular sensing, and chirality tuning are discussed, and a perspective of emerging concepts such as Pancharatnam−Berry (PB) phase in this booming research field is presented.
DOI
The plasmonic chiroptical effect has been used to manipulate chiral states of light, where the strong field enhancement and light localization in metallic nanostructures can amplify the chiroptical response. Moreover, in metamaterials, the chiroptical effect leads to circular dichroism (CD), circular birefringence (CB), and asymmetric transmission. Potential applications enabled by chiral plasmonics have been realized in various areas of nanoscience and nanotechnology. In this review, both basic theories and state-of-the-art studies on plasmonic chiroptical effects are summarized. Molecular chiroptical effects are drastically enhanced by metallic nanostructures that can generate a “superchiral” field, which arises from the strong electromagnetic interactions. Both intrinsic and extrinsic plasmonic chiral metamaterials formed by the periodic arrangement of metallic nanostructured units show high levels of CB, CD, and asymmetric transmission. Consequent applications including photo detection, molecular sensing, and chirality tuning are discussed, and a perspective of emerging concepts such as Pancharatnam−Berry (PB) phase in this booming research field is presented.
DOI
Wednesday, June 28, 2017
Temperature-induced Coalescence of Droplets Manipulated by Optical Trapping in an Oil-in-Water Emulsion
Manami MITSUNOBU, Sakurako KOBAYASHI, Nobuyuki TAKEYASU, Takashi KANETA
Coalescence of oil droplets in an oil-in-water (O/W) emulsion was achieved with heating and optical trapping. Three types of O/W emulsions were prepared by adding a mixture of butanol and n-decane to an aqueous solution containing a cationic surfactant (cetyltrimethylammonium bromide, CTAB), an anionic surfactant (sodium dodecyl sulfate, SDS), or a neutral hydrophilic polymer (polyethylene glycol, PEG) as an emulsifier. Two oil droplets in the emulsions were randomly trapped in a square capillary tube by two laser beams in order to induce coalescence. Coalescence of the droplets could not be achieved at room temperature (25°C) regardless of the type of emulsifier. Conversely, the droplets prepared with PEG coalesced at a temperature higher than 30°C, although the droplets with ionic surfactants CTAB and SDS did not coalesce even at the elevated temperature due to their electrostatic repulsion. The size of the resultant coalesced droplet was consistent with that calculated from the size of the two droplets of oil, which indicated successful coalescence of the two droplets. We also found that the time required for the coalescence could be correlated with the temperature using an Arrhenius plot.
DOI
Coalescence of oil droplets in an oil-in-water (O/W) emulsion was achieved with heating and optical trapping. Three types of O/W emulsions were prepared by adding a mixture of butanol and n-decane to an aqueous solution containing a cationic surfactant (cetyltrimethylammonium bromide, CTAB), an anionic surfactant (sodium dodecyl sulfate, SDS), or a neutral hydrophilic polymer (polyethylene glycol, PEG) as an emulsifier. Two oil droplets in the emulsions were randomly trapped in a square capillary tube by two laser beams in order to induce coalescence. Coalescence of the droplets could not be achieved at room temperature (25°C) regardless of the type of emulsifier. Conversely, the droplets prepared with PEG coalesced at a temperature higher than 30°C, although the droplets with ionic surfactants CTAB and SDS did not coalesce even at the elevated temperature due to their electrostatic repulsion. The size of the resultant coalesced droplet was consistent with that calculated from the size of the two droplets of oil, which indicated successful coalescence of the two droplets. We also found that the time required for the coalescence could be correlated with the temperature using an Arrhenius plot.
DOI
Vector assembly of colloids on monolayer substrates
Lingxiang Jiang, Shenyu Yang, Boyce Tsang, Mei Tu & Steve Granick
The key to spontaneous and directed assembly is to encode the desired assembly information to building blocks in a programmable and efficient way. In computer graphics, raster graphics encodes images on a single-pixel level, conferring fine details at the expense of large file sizes, whereas vector graphics encrypts shape information into vectors that allow small file sizes and operational transformations. Here, we adapt this raster/vector concept to a 2D colloidal system and realize ‘vector assembly’ by manipulating particles on a colloidal monolayer substrate with optical tweezers. In contrast to raster assembly that assigns optical tweezers to each particle, vector assembly requires a minimal number of optical tweezers that allow operations like chain elongation and shortening. This vector approach enables simple uniform particles to form a vast collection of colloidal arenes and colloidenes, the spontaneous dissociation of which is achieved with precision and stage-by-stage complexity by simply removing the optical tweezers.
DOI
The key to spontaneous and directed assembly is to encode the desired assembly information to building blocks in a programmable and efficient way. In computer graphics, raster graphics encodes images on a single-pixel level, conferring fine details at the expense of large file sizes, whereas vector graphics encrypts shape information into vectors that allow small file sizes and operational transformations. Here, we adapt this raster/vector concept to a 2D colloidal system and realize ‘vector assembly’ by manipulating particles on a colloidal monolayer substrate with optical tweezers. In contrast to raster assembly that assigns optical tweezers to each particle, vector assembly requires a minimal number of optical tweezers that allow operations like chain elongation and shortening. This vector approach enables simple uniform particles to form a vast collection of colloidal arenes and colloidenes, the spontaneous dissociation of which is achieved with precision and stage-by-stage complexity by simply removing the optical tweezers.
DOI
Propagation properties and radiation forces of the Airy Gaussian vortex beams in a harmonic potential
Zihao Pang and Dongmei Deng
We investigate the propagation properties and the radiation forces of Airy Gaussian vortex (AiGV) beams in a harmonic potential analytically and numerically in this paper. Obtaining the propagation expression of AiGV beams by solving the dimensionless linear (2+1) D Schrödinger equation in a harmonic potential, we perform the track, the intensity and phase distributions, the propagation shapes, the energy flow and the angular momentum of AiGV beams in a harmonic potential with the method of numerical simulations. The trajectory acting like a cosine curve is shown. Periodic inversion and phase oscillation are demonstrated during propagation. The influence of the distribution factor and the vortex factor on the propagation of AiGV beams in a harmonic potential are discussed. Likewise, the motion of the Poynting vector and the angular momentum is elucidated respectively. As for the radiation forces, we explore the gradient and scattering forces on Rayleigh dielectric particles induced by AiGV beams. In particular, it’s found that the value of the scattering force is approximately seven orders of magnitude larger than that of the gradient force during the propagation in a harmonic potential.
DOI
We investigate the propagation properties and the radiation forces of Airy Gaussian vortex (AiGV) beams in a harmonic potential analytically and numerically in this paper. Obtaining the propagation expression of AiGV beams by solving the dimensionless linear (2+1) D Schrödinger equation in a harmonic potential, we perform the track, the intensity and phase distributions, the propagation shapes, the energy flow and the angular momentum of AiGV beams in a harmonic potential with the method of numerical simulations. The trajectory acting like a cosine curve is shown. Periodic inversion and phase oscillation are demonstrated during propagation. The influence of the distribution factor and the vortex factor on the propagation of AiGV beams in a harmonic potential are discussed. Likewise, the motion of the Poynting vector and the angular momentum is elucidated respectively. As for the radiation forces, we explore the gradient and scattering forces on Rayleigh dielectric particles induced by AiGV beams. In particular, it’s found that the value of the scattering force is approximately seven orders of magnitude larger than that of the gradient force during the propagation in a harmonic potential.
DOI
Plasmonic optical trapping of nanometer-sized J- /H- dye aggregates as explored by fluorescence microspectroscopy
Ayaka Mototsuji, Tatsuya Shoji, Yumi Wakisaka, Kei Murakoshi, Hiroshi Yao, and Yasuyuki Tsuboi
In the present study, we explored plasmonic optical trapping (POT) of nanometer-sized organic crystals, carbocyanine dye aggregates (JC-1). JC-1 dye forms both J- and H- aggregates in aqueous solution. POT behavior was analyzed using fluorescence microspectroscopy. POT of JC-1 aggregates was realized in an increase in their fluorescence intensity from the focus area upon plasmon excitation. Repeating on-and-off plasmonic excitation resulted in POT of JC-1 aggregates in a trap-and-release mode. Such POT of nanometer-sized dye aggregates lying in a Rayleigh scattering regime (< 100 nm) is important toward molecular manipulation. Furthermore, interestingly, we found that the J-aggregates were preferentially trapped than H-aggregates. It possibly indicates semi-selective optical trapping of nanoparticles on the basis of molecular alignments.
DOI
In the present study, we explored plasmonic optical trapping (POT) of nanometer-sized organic crystals, carbocyanine dye aggregates (JC-1). JC-1 dye forms both J- and H- aggregates in aqueous solution. POT behavior was analyzed using fluorescence microspectroscopy. POT of JC-1 aggregates was realized in an increase in their fluorescence intensity from the focus area upon plasmon excitation. Repeating on-and-off plasmonic excitation resulted in POT of JC-1 aggregates in a trap-and-release mode. Such POT of nanometer-sized dye aggregates lying in a Rayleigh scattering regime (< 100 nm) is important toward molecular manipulation. Furthermore, interestingly, we found that the J-aggregates were preferentially trapped than H-aggregates. It possibly indicates semi-selective optical trapping of nanoparticles on the basis of molecular alignments.
DOI
High-resolution and multi-range particle separation by microscopic vibration in an optofluidic chip
Y. Z. Shi, S. Xiong, L. K. Chin, Y. Yang, J. B. Zhang, W. Ser, J. H. Wu, T. N. Chen, Z. C. Yang, Y. L. Hao, B. Liedberg, P. H. Yap, Y. Zhang and A. Q. Liu
An optofluidic chip is demonstrated in experiments for high-resolution and multi-range particle separation through the optically-induced microscopic vibration effect, where nanoparticles are trapped in loosely overdamped optical potential wells created with combined optical and fluidic constraints. It is the first demonstration of separating single nanoparticles with diameters ranging from 60 to 100 nm with a resolution of 10 nm. Nanoparticles vibrate with an amplitude of 3–7 μm in the loosely overdamped potential wells in the microchannel. The proposed optofluidic device is capable of high-resolution particle separation at both nanoscale and microscale without reconfiguring the device. The separation of bacteria from other larger cells is accomplished using the same chip and operation conditions. The unique trapping mechanism and the superb performance in high-resolution and multi-range particle separation of the proposed optofluidic chip promise great potential for a diverse range of biomedical applications.
DOI
An optofluidic chip is demonstrated in experiments for high-resolution and multi-range particle separation through the optically-induced microscopic vibration effect, where nanoparticles are trapped in loosely overdamped optical potential wells created with combined optical and fluidic constraints. It is the first demonstration of separating single nanoparticles with diameters ranging from 60 to 100 nm with a resolution of 10 nm. Nanoparticles vibrate with an amplitude of 3–7 μm in the loosely overdamped potential wells in the microchannel. The proposed optofluidic device is capable of high-resolution particle separation at both nanoscale and microscale without reconfiguring the device. The separation of bacteria from other larger cells is accomplished using the same chip and operation conditions. The unique trapping mechanism and the superb performance in high-resolution and multi-range particle separation of the proposed optofluidic chip promise great potential for a diverse range of biomedical applications.
DOI
Direct measurement of optical trapping force gradient on polystyrene microspheres using a carbon nanotube mechanical resonator
Masaaki Yasuda, Kuniharu Takei, Takayuki Arie & Seiji Akita
Optical tweezers based on optical radiation pressure are widely used to manipulate nanoscale to microscale particles. This study demonstrates direct measurement of the optical force gradient distribution acting on a polystyrene (PS) microsphere using a carbon nanotube (CNT) mechanical resonator, where a PS microsphere with 3 μm diameter is welded at the CNT tip using laser heating. With the CNT mechanical resonator with PS microsphere, we measured the distribution of optical force gradient with resolution near the thermal noise limit of 0.02 pN/μm in vacuum, in which condition enables us to high accuracy measurement using the CNT mechanical resonator because of reduced mechanical damping from surrounding fluid. The obtained force gradient and the force gradient distribution agree well with theoretical values calculated using Lorenz–Mie theory.
DOI
Optical tweezers based on optical radiation pressure are widely used to manipulate nanoscale to microscale particles. This study demonstrates direct measurement of the optical force gradient distribution acting on a polystyrene (PS) microsphere using a carbon nanotube (CNT) mechanical resonator, where a PS microsphere with 3 μm diameter is welded at the CNT tip using laser heating. With the CNT mechanical resonator with PS microsphere, we measured the distribution of optical force gradient with resolution near the thermal noise limit of 0.02 pN/μm in vacuum, in which condition enables us to high accuracy measurement using the CNT mechanical resonator because of reduced mechanical damping from surrounding fluid. The obtained force gradient and the force gradient distribution agree well with theoretical values calculated using Lorenz–Mie theory.
DOI
Tuesday, June 27, 2017
Metasurfaces and Colloidal Suspensions Composed of 3D Chiral Si Nanoresonators
Ruggero Verre, Lei Shao, Nils Odebo Länk, Pawel Karpinski, Andrew B. Yankovich, Tomasz J. Antosiewicz, Eva Olsson, Mikael Käll
High-refractive-index silicon nanoresonators are promising low-loss alternatives to plasmonic particles in CMOS-compatible nanophotonics applications. However, complex 3D particle morphologies are challenging to realize in practice, thus limiting the range of achievable optical functionalities. Using 3D film structuring and a novel gradient mask transfer technique, the first intrinsically chiral dielectric metasurface is fabricated in the form of a monolayer of twisted silicon nanocrescents that can be easily detached and dissolved into colloidal suspension. The metasurfaces exhibit selective handedness and a circular dichroism as large as 160° µm−1 due to pronounced differences in induced current loops for left-handed and right-handed polarization. The detailed morphology of the detached particles is analyzed using high-resolution transmission electron microscopy. Furthermore, it is shown that the particles can be manipulated in solution using optical tweezers. The fabrication and detachment method can be extended to different nanoparticle geometries and paves the way for a wide range of novel nanophotonic experiments and applications of high-index dielectrics.
DOI
High-refractive-index silicon nanoresonators are promising low-loss alternatives to plasmonic particles in CMOS-compatible nanophotonics applications. However, complex 3D particle morphologies are challenging to realize in practice, thus limiting the range of achievable optical functionalities. Using 3D film structuring and a novel gradient mask transfer technique, the first intrinsically chiral dielectric metasurface is fabricated in the form of a monolayer of twisted silicon nanocrescents that can be easily detached and dissolved into colloidal suspension. The metasurfaces exhibit selective handedness and a circular dichroism as large as 160° µm−1 due to pronounced differences in induced current loops for left-handed and right-handed polarization. The detailed morphology of the detached particles is analyzed using high-resolution transmission electron microscopy. Furthermore, it is shown that the particles can be manipulated in solution using optical tweezers. The fabrication and detachment method can be extended to different nanoparticle geometries and paves the way for a wide range of novel nanophotonic experiments and applications of high-index dielectrics.
DOI
Biolens behavior of RBCs under optically-induced mechanical stress
Francesco Merola, Álvaro Barroso, Lisa Miccio, Pasquale Memmolo, Martina Mugnano, Pietro Ferraro, Cornelia Denz
In this work, the optical behavior of Red Blood Cells (RBCs) under an optically-induced mechanical stress was studied. Exploiting the new findings concerning the optical lens-like behavior of RBCs, the variations of the wavefront refracted by optically-deformed RBCs were further investigated. Experimental analysis have been performed through the combination of digital holography and numerical analysis based on Zernike polynomials, while the biological lens is deformed under the action of multiple dynamic optical tweezers. Detailed wavefront analysis provides comprehensive information about the aberrations induced by the applied mechanical stress. By this approach it was shown that the optical properties of RBCs in their discocyte form can be affected in a different way depending on the geometry of the deformation. In analogy to classical optical testing procedures, optical parameters can be correlated to a particular mechanical deformation. This could open new routes for analyzing cell elasticity by examining optical parameters instead of direct but with low resolution strain analysis, thanks to the high sensitivity of the interferometric tool. Future application of this approach could lead to early detection and diagnosis of blood diseases through a single-step wavefront analysis for evaluating different cells elasticity.
DOI
In this work, the optical behavior of Red Blood Cells (RBCs) under an optically-induced mechanical stress was studied. Exploiting the new findings concerning the optical lens-like behavior of RBCs, the variations of the wavefront refracted by optically-deformed RBCs were further investigated. Experimental analysis have been performed through the combination of digital holography and numerical analysis based on Zernike polynomials, while the biological lens is deformed under the action of multiple dynamic optical tweezers. Detailed wavefront analysis provides comprehensive information about the aberrations induced by the applied mechanical stress. By this approach it was shown that the optical properties of RBCs in their discocyte form can be affected in a different way depending on the geometry of the deformation. In analogy to classical optical testing procedures, optical parameters can be correlated to a particular mechanical deformation. This could open new routes for analyzing cell elasticity by examining optical parameters instead of direct but with low resolution strain analysis, thanks to the high sensitivity of the interferometric tool. Future application of this approach could lead to early detection and diagnosis of blood diseases through a single-step wavefront analysis for evaluating different cells elasticity.
DOI
β -Cyclodextrin polymer binding to DNA: Modulating the physicochemical parameters
J. C. B. Rocha, E. F. Silva, M. F. Oliveira, F. B. Sousa, A. V. N. C. Teixeira, and M. S. Rocha
Cyclodextrins and cyclodextrins-modified molecules have interesting and appealing properties due to their capacity to host components that are normally insoluble or poorly soluble in water. In this work, we investigate the interaction of a β-cyclodextrin polymer (poly-β-CD) with λ-DNA. The polymers are obtained by the reaction of β-CD with epichlorohydrin in alkaline conditions. We have used optical tweezers to characterize the changes of the mechanical properties of DNA molecules by increasing the concentration of poly-β-CD in the sample. The physical chemistry of the interaction is then deduced from these measurements by using a recently developed quenched-disorder statistical model. It is shown that the contour length of the DNA does not change in the whole range of poly-β-CD concentration (<300μM). On the other hand, significant alterations were observed in the persistence length that identifies two binding modes corresponding to the clustering of ∼2.6 and ∼14 polymer molecules along the DNA double helix, depending on the polymer concentration. Comparing these results with the ones obtained for monomeric β-CD, it was observed that the concentration of CD that alters the DNA persistence length is considerably smaller when in the polymeric form. Also, the binding constant of the polymer-DNA interaction is three orders of magnitude higher than the one found for native (monomeric) β-CD. These results show that the polymerization of the β-CD strongly increases its binding affinity to the DNA molecule. This property can be wisely used to modulate the binding of cyclodextrins to the DNA double helix.
DOI
Cyclodextrins and cyclodextrins-modified molecules have interesting and appealing properties due to their capacity to host components that are normally insoluble or poorly soluble in water. In this work, we investigate the interaction of a β-cyclodextrin polymer (poly-β-CD) with λ-DNA. The polymers are obtained by the reaction of β-CD with epichlorohydrin in alkaline conditions. We have used optical tweezers to characterize the changes of the mechanical properties of DNA molecules by increasing the concentration of poly-β-CD in the sample. The physical chemistry of the interaction is then deduced from these measurements by using a recently developed quenched-disorder statistical model. It is shown that the contour length of the DNA does not change in the whole range of poly-β-CD concentration (<300μM). On the other hand, significant alterations were observed in the persistence length that identifies two binding modes corresponding to the clustering of ∼2.6 and ∼14 polymer molecules along the DNA double helix, depending on the polymer concentration. Comparing these results with the ones obtained for monomeric β-CD, it was observed that the concentration of CD that alters the DNA persistence length is considerably smaller when in the polymeric form. Also, the binding constant of the polymer-DNA interaction is three orders of magnitude higher than the one found for native (monomeric) β-CD. These results show that the polymerization of the β-CD strongly increases its binding affinity to the DNA molecule. This property can be wisely used to modulate the binding of cyclodextrins to the DNA double helix.
DOI
Experimental comparison of forces resisting viral DNA packaging and driving DNA ejection
Nicholas Keller, Zachary T. Berndsen, Paul J. Jardine, and Douglas E. Smith
We compare forces resisting DNA packaging and forces driving DNA ejection in bacteriophage phi29 with theoretical predictions. Ejection of DNA from prohead-motor complexes is triggered by heating complexes after in vitro packaging and force is inferred from the suppression of ejection by applied osmotic pressure. Ejection force from 0% to 80% filling is found to be in quantitative agreement with predictions of a continuum mechanics model that assumes a repulsive DNA-DNA interaction potential based on DNA condensation studies and predicts an inverse-spool conformation. Force resisting DNA packaging from ∼80% to 100% filling inferred from optical tweezers studies is also consistent with the predictions of this model. The striking agreement with these two different measurements suggests that the overall energetics of DNA packaging is well described by the model. However, since electron microscopy studies of phi29 do not reveal a spool conformation, our findings suggest that the spool model overestimates the role of bending rigidity and underestimates the role of intrastrand repulsion. Below ∼80% filling the inferred forces resisting packaging are unexpectedly lower than the inferred ejection forces, suggesting that in this filling range the forces are less accurately determined or strongly temperature dependent.
DOI
We compare forces resisting DNA packaging and forces driving DNA ejection in bacteriophage phi29 with theoretical predictions. Ejection of DNA from prohead-motor complexes is triggered by heating complexes after in vitro packaging and force is inferred from the suppression of ejection by applied osmotic pressure. Ejection force from 0% to 80% filling is found to be in quantitative agreement with predictions of a continuum mechanics model that assumes a repulsive DNA-DNA interaction potential based on DNA condensation studies and predicts an inverse-spool conformation. Force resisting DNA packaging from ∼80% to 100% filling inferred from optical tweezers studies is also consistent with the predictions of this model. The striking agreement with these two different measurements suggests that the overall energetics of DNA packaging is well described by the model. However, since electron microscopy studies of phi29 do not reveal a spool conformation, our findings suggest that the spool model overestimates the role of bending rigidity and underestimates the role of intrastrand repulsion. Below ∼80% filling the inferred forces resisting packaging are unexpectedly lower than the inferred ejection forces, suggesting that in this filling range the forces are less accurately determined or strongly temperature dependent.
DOI
A unique profilin-actin interface is important for malaria parasite motility
Catherine A. Moreau, Saligram P. Bhargav , Hirdesh Kumar , Katharina A. Quadt , Henni Piirainen, Léanne Strauss, Jessica Kehrer, Martin Streichfuss, Joachim P. Spatz, Rebecca C. Wade, Inari Kursula , Friedrich Frischknecht
Profilin is an actin monomer binding protein that provides ATP-actin for incorporation into actin filaments. In contrast to higher eukaryotic cells with their large filamentous actin structures, apicomplexan parasites typically contain only short and highly dynamic microfilaments. In apicomplexans, profilin appears to be the main monomer-sequestering protein. Compared to classical profilins, apicomplexan profilins contain an additional arm-like β-hairpin motif, which we show here to be critically involved in actin binding. Through comparative analysis using two profilin mutants, we reveal this motif to be implicated in gliding motility of Plasmodium berghei sporozoites, the rapidly migrating forms of a rodent malaria parasite transmitted by mosquitoes. Force measurements on migrating sporozoites and molecular dynamics simulations indicate that the interaction between actin and profilin fine-tunes gliding motility. Our data suggest that evolutionary pressure to achieve efficient high-speed gliding has resulted in a unique profilin-actin interface in these parasites.
DOI
Profilin is an actin monomer binding protein that provides ATP-actin for incorporation into actin filaments. In contrast to higher eukaryotic cells with their large filamentous actin structures, apicomplexan parasites typically contain only short and highly dynamic microfilaments. In apicomplexans, profilin appears to be the main monomer-sequestering protein. Compared to classical profilins, apicomplexan profilins contain an additional arm-like β-hairpin motif, which we show here to be critically involved in actin binding. Through comparative analysis using two profilin mutants, we reveal this motif to be implicated in gliding motility of Plasmodium berghei sporozoites, the rapidly migrating forms of a rodent malaria parasite transmitted by mosquitoes. Force measurements on migrating sporozoites and molecular dynamics simulations indicate that the interaction between actin and profilin fine-tunes gliding motility. Our data suggest that evolutionary pressure to achieve efficient high-speed gliding has resulted in a unique profilin-actin interface in these parasites.
DOI
Trapping Two Types of Particles Using a Laguerre–Gaussian Correlated Schell-Model Beam
Yuan Zhou; Hua-Feng Xu; Yangsheng Yuan; Ji Peng; Jun Qu; Wei Huang
Based on the Rayleigh scattering theory, the radiation forces and the trap stiffness on Rayleigh dielectric sphere induced by a focused Laguerre-Gaussian correlated Schell-model (LGCSM) beam are theoretically studied. It is found that by choosing the appropriate transverse coherence width, mode orders, transverse beam width, and focus lengths, a Rayleigh particle whose refractive index is larger or smaller than the ambient medium can be trapped. Our results will have some theoretical reference value for optical trapping.
DOI
Based on the Rayleigh scattering theory, the radiation forces and the trap stiffness on Rayleigh dielectric sphere induced by a focused Laguerre-Gaussian correlated Schell-model (LGCSM) beam are theoretically studied. It is found that by choosing the appropriate transverse coherence width, mode orders, transverse beam width, and focus lengths, a Rayleigh particle whose refractive index is larger or smaller than the ambient medium can be trapped. Our results will have some theoretical reference value for optical trapping.
DOI
Friday, June 23, 2017
Transition Path Times Measured by Single-Molecule Spectroscopy
Hoi Sung Chung
The transition path is a tiny fraction of a molecular trajectory during which the free-energy barrier is crossed. It is a single-molecule property and contains all mechanistic information of folding processes of biomolecules such as proteins and nucleic acids. However, the transition path has been difficult to probe because it is short and rarely visited when transitions actually occur. Recent technical advances in single-molecule spectroscopy have made it possible to directly probe transition paths, which has opened up new theoretical and experimental approaches to investigating folding mechanisms. This article reviews recent single-molecule fluorescence and force spectroscopic measurements of transition path times and their connection to both theory and simulations.
DOI
The transition path is a tiny fraction of a molecular trajectory during which the free-energy barrier is crossed. It is a single-molecule property and contains all mechanistic information of folding processes of biomolecules such as proteins and nucleic acids. However, the transition path has been difficult to probe because it is short and rarely visited when transitions actually occur. Recent technical advances in single-molecule spectroscopy have made it possible to directly probe transition paths, which has opened up new theoretical and experimental approaches to investigating folding mechanisms. This article reviews recent single-molecule fluorescence and force spectroscopic measurements of transition path times and their connection to both theory and simulations.
DOI
Combined single molecule experimental and computational approaches for understanding the unfolding pathway of a viral translation enhancer that participates in a conformational switch
My-Tra Le, Wojciech K Kasprzak, Bruce A. Shapiro & Anne E Simon
How plus-strand [+]RNA virus genomes transition from translation templates to replication templates is a matter of much speculation. We have previously proposed that, for Turnip crinkle virus, binding of the encoded RNA-dependent RNA polymerase (RdRp) to the 3′UTR of the [+]RNA template promotes a regional wide-spread conformational switch to an alternative structure that disassembles the cap-independent translation element (CITE) in the 3′UTR. The active 3′CITE folds into a tRNA-like T-shaped structure (TSS) that binds to 80S ribosomes and 60S subunits in the P-site. In this Point-of-View, we discuss the history of our research on the TSS and our recent report combining coarse level single molecule force spectroscopy (optical tweezers) with fine-grain computer simulations of this experimental process and biochemical approaches to obtain a detailed understanding of how RdRp binding in the TSS vicinity might lead to an extensive rearrangement of the RNA structure.
DOI
How plus-strand [+]RNA virus genomes transition from translation templates to replication templates is a matter of much speculation. We have previously proposed that, for Turnip crinkle virus, binding of the encoded RNA-dependent RNA polymerase (RdRp) to the 3′UTR of the [+]RNA template promotes a regional wide-spread conformational switch to an alternative structure that disassembles the cap-independent translation element (CITE) in the 3′UTR. The active 3′CITE folds into a tRNA-like T-shaped structure (TSS) that binds to 80S ribosomes and 60S subunits in the P-site. In this Point-of-View, we discuss the history of our research on the TSS and our recent report combining coarse level single molecule force spectroscopy (optical tweezers) with fine-grain computer simulations of this experimental process and biochemical approaches to obtain a detailed understanding of how RdRp binding in the TSS vicinity might lead to an extensive rearrangement of the RNA structure.
DOI
Cooperativity of myosin interaction with thin filaments is enhanced by stabilizing substitutions in tropomyosin
Daniil V. Shchepkin, Salavat R. Nabiev, Galina V. Kopylova, Alexander M. Matyushenko, Dmitrii I. Levitsky, Sergey Y. Bershitsky, Andrey K. Tsaturyan
Muscle contraction is powered by myosin interaction with actin-based thin filaments containing Ca2+-regulatory proteins, tropomyosin and troponin. Coiled-coil tropomyosin molecules form a long helical strand that winds around actin filament and either shields actin from myosin binding or opens it. Non-canonical residues G126 and D137 in the central part of tropomyosin destabilize its coiled-coil structure. Their substitutions for canonical ones, G126R and D137L, increase structural stability and the velocity of sliding of reconstructed thin filaments along myosin coated surface. The effect of these stabilizing mutations on force of the actin–myosin interaction is unknown. It also remains unclear whether the stabilization affects single actin–myosin interactions or it modifies the cooperativity of the binding of myosin molecules to actin. We used an optical trap to measure the effects of the stabilization on step size, unitary force and duration of the interactions at low and high load and compared the results with those obtained in an in vitro motility assay. We found that significant prolongation of lifetime of the actin–myosin complex under high load observed at high extent of tropomyosin stabilization, i.e. with double mutant, G126R/D137L, correlates with higher force in the motility assay. Also, the higher the extent of stabilization of tropomyosin, the fewer myosin molecules are needed to propel the thin filaments. The data suggest that the effects of the stabilizing mutations in tropomyosin on the myosin interaction with regulated thin filaments are mainly realized via cooperative mechanisms by increasing the size of cooperative unit.
DOI
Muscle contraction is powered by myosin interaction with actin-based thin filaments containing Ca2+-regulatory proteins, tropomyosin and troponin. Coiled-coil tropomyosin molecules form a long helical strand that winds around actin filament and either shields actin from myosin binding or opens it. Non-canonical residues G126 and D137 in the central part of tropomyosin destabilize its coiled-coil structure. Their substitutions for canonical ones, G126R and D137L, increase structural stability and the velocity of sliding of reconstructed thin filaments along myosin coated surface. The effect of these stabilizing mutations on force of the actin–myosin interaction is unknown. It also remains unclear whether the stabilization affects single actin–myosin interactions or it modifies the cooperativity of the binding of myosin molecules to actin. We used an optical trap to measure the effects of the stabilization on step size, unitary force and duration of the interactions at low and high load and compared the results with those obtained in an in vitro motility assay. We found that significant prolongation of lifetime of the actin–myosin complex under high load observed at high extent of tropomyosin stabilization, i.e. with double mutant, G126R/D137L, correlates with higher force in the motility assay. Also, the higher the extent of stabilization of tropomyosin, the fewer myosin molecules are needed to propel the thin filaments. The data suggest that the effects of the stabilizing mutations in tropomyosin on the myosin interaction with regulated thin filaments are mainly realized via cooperative mechanisms by increasing the size of cooperative unit.
DOI
Setting up of holographic optical tweezer arrays
Deepak K. Gupta, B. V. R. Tata, and T. R. Ravindran
Optical tweezers use tightly focused laser beams to hold and move microscopic objects in a solvent. However, many applications require simultaneous control over multitude of particles, positioning them in 3D space at desired locations with desired symmetry, which is made possible by the use of holographic optical tweezers using the technique of beam shaping and holography. We have designed and developed a holographic optical tweezer set-up using a phase only liquid crystal, reflective spatial light modulator. We employ the technique of phase modulation to modulate the phase of the beam by generating holograms using Random Superposition (RS) and weighted Gerchberg Saxton algorithm (WGS) algorithm for generating desired patterns of light at the trapping plane. A 4×4 array of beams with square symmetry was generated using WGS algorithm and trapped polystyrene particles of size 1.2 micron in a 4×4 two dimensional array. There were uniformity issues among the trap intensities, as we move away from the zeroth order spot. This was corrected by taking into account diffraction effects due to the pixelated nature of SLM modulating the intensity of the trap spots and the ghost order suppression by spatial disorder.
DOI
Optical tweezers use tightly focused laser beams to hold and move microscopic objects in a solvent. However, many applications require simultaneous control over multitude of particles, positioning them in 3D space at desired locations with desired symmetry, which is made possible by the use of holographic optical tweezers using the technique of beam shaping and holography. We have designed and developed a holographic optical tweezer set-up using a phase only liquid crystal, reflective spatial light modulator. We employ the technique of phase modulation to modulate the phase of the beam by generating holograms using Random Superposition (RS) and weighted Gerchberg Saxton algorithm (WGS) algorithm for generating desired patterns of light at the trapping plane. A 4×4 array of beams with square symmetry was generated using WGS algorithm and trapped polystyrene particles of size 1.2 micron in a 4×4 two dimensional array. There were uniformity issues among the trap intensities, as we move away from the zeroth order spot. This was corrected by taking into account diffraction effects due to the pixelated nature of SLM modulating the intensity of the trap spots and the ghost order suppression by spatial disorder.
DOI
Holographic fluorescence mapping using space-division matching method
Ryosuke Abe, Yoshio Hayasaki
Three-dimensional mapping of fluorescence light sources was performed by using self-interference digital holography. The positions of the sources were quantitatively determined by using Gaussian fitting of the axial and lateral intensity distributions obtained from diffraction calculations through position calibration from the observation space to the sample space. A space-division matching method was developed to perform the mapping of many fluorescence light sources, in this experiment, 500 nm fluorescent nanoparticles fixed in gelatin. A fluorescence digital holographic microscope having a 60× objective lens with a numerical aperture of 1.25 detected 13 fluorescence light sources in a measurable region with a radius of View the MathML source and a height of View the MathML source. It was found that the measurable region had a conical shape resulting from the overlap between two beams.
DOI
Three-dimensional mapping of fluorescence light sources was performed by using self-interference digital holography. The positions of the sources were quantitatively determined by using Gaussian fitting of the axial and lateral intensity distributions obtained from diffraction calculations through position calibration from the observation space to the sample space. A space-division matching method was developed to perform the mapping of many fluorescence light sources, in this experiment, 500 nm fluorescent nanoparticles fixed in gelatin. A fluorescence digital holographic microscope having a 60× objective lens with a numerical aperture of 1.25 detected 13 fluorescence light sources in a measurable region with a radius of View the MathML source and a height of View the MathML source. It was found that the measurable region had a conical shape resulting from the overlap between two beams.
DOI
Tuesday, June 20, 2017
Nanofibre optic force transducers with sub-piconewton resolution via near-field plasmon–dielectric interactions
Qian Huang, Joon Lee, Fernando Teran Arce, Ilsun Yoon, Pavimol Angsantikul, Justin Liu, Yuesong Shi, Josh Villanueva, Soracha Thamphiwatana, Xuanyi Ma, Liangfang Zhang, Shaochen Chen, Ratnesh Lal & Donald J. Sirbuly
Ultrasensitive nanomechanical instruments, including the atomic force microscope (AFM) and optical and magnetic tweezers have helped shed new light on the complex mechanical environments of biological processes. However, it is difficult to scale down the size of these instruments due to their feedback mechanisms, which, if overcome, would enable high-density nanomechanical probing inside materials. A variety of molecular force probes including mechanophores, quantum dots, fluorescent pairs and molecular rotors have been designed to measure intracellular stresses; however, fluorescence-based techniques can have short operating times due to photo-instability and it is still challenging to quantify the forces with high spatial and mechanical resolution. Here, we develop a compact nanofibre optic force transducer (NOFT) that utilizes strong near-field plasmon–dielectric interactions to measure local forces with a sensitivity of <200 fN. The NOFT system is tested by monitoring bacterial motion and heart-cell beating as well as detecting infrasound power in solution.
DOI
Ultrasensitive nanomechanical instruments, including the atomic force microscope (AFM) and optical and magnetic tweezers have helped shed new light on the complex mechanical environments of biological processes. However, it is difficult to scale down the size of these instruments due to their feedback mechanisms, which, if overcome, would enable high-density nanomechanical probing inside materials. A variety of molecular force probes including mechanophores, quantum dots, fluorescent pairs and molecular rotors have been designed to measure intracellular stresses; however, fluorescence-based techniques can have short operating times due to photo-instability and it is still challenging to quantify the forces with high spatial and mechanical resolution. Here, we develop a compact nanofibre optic force transducer (NOFT) that utilizes strong near-field plasmon–dielectric interactions to measure local forces with a sensitivity of <200 fN. The NOFT system is tested by monitoring bacterial motion and heart-cell beating as well as detecting infrasound power in solution.
DOI
Dynamic chromatin technologies: from individual molecules to epigenomic regulation in cells
Olivier Cuvier & Beat Fierz
The establishment and maintenance of chromatin states involves multiscale dynamic processes integrating transcription factor and multiprotein effector dynamics, cycles of chemical chromatin modifications, and chromatin structural organization. Recent developments in genomic technologies are emerging that are enabling a view beyond ensemble- and time-averaged properties and are revealing the importance of dynamic chromatin states for cell fate decisions, differentiation and reprogramming at the single-cell level. Concurrently, biochemical and single-molecule methodologies are providing key insights into the underlying molecular mechanisms. Combining results from defined in vitro and single-molecule studies with single-cell genomic approaches thus holds great promise for understanding chromatin-based transcriptional memory and cell fate. In this Review, we discuss recent developments in biochemical, single-molecule biophysical and single-cell genomic technologies and review how the findings from these approaches can be integrated to paint a comprehensive picture of dynamic chromatin states.
DOI
The establishment and maintenance of chromatin states involves multiscale dynamic processes integrating transcription factor and multiprotein effector dynamics, cycles of chemical chromatin modifications, and chromatin structural organization. Recent developments in genomic technologies are emerging that are enabling a view beyond ensemble- and time-averaged properties and are revealing the importance of dynamic chromatin states for cell fate decisions, differentiation and reprogramming at the single-cell level. Concurrently, biochemical and single-molecule methodologies are providing key insights into the underlying molecular mechanisms. Combining results from defined in vitro and single-molecule studies with single-cell genomic approaches thus holds great promise for understanding chromatin-based transcriptional memory and cell fate. In this Review, we discuss recent developments in biochemical, single-molecule biophysical and single-cell genomic technologies and review how the findings from these approaches can be integrated to paint a comprehensive picture of dynamic chromatin states.
DOI
Mechanism of ribosome translation through mRNA secondary structures
Ping Xie, Hong Chen
A ribosome is a macromolecular machine that is responsible for translating the genetic codes in messenger RNA (mRNA) into polypeptide chains. It has been determined that besides translating through the single-stranded region, the ribosome can also translate through the duplex region of mRNA by unwinding the duplex. To understand the mechanism of ribosome translation through the duplex, several models have been proposed to study the dynamics of mRNA unwinding. Here, we present a comprehensive review of these models and also discuss other possible models. We evaluate each model and discuss the consistency and/or inconsistency between the theoretical results that are obtained based on each model and the available experimental data, thus determining which model is the most reasonable one to describe the mRNA unwinding mechanism and dynamics of the ribosome. Moreover, a framework for future studies in this subject is provided.
DOI
A ribosome is a macromolecular machine that is responsible for translating the genetic codes in messenger RNA (mRNA) into polypeptide chains. It has been determined that besides translating through the single-stranded region, the ribosome can also translate through the duplex region of mRNA by unwinding the duplex. To understand the mechanism of ribosome translation through the duplex, several models have been proposed to study the dynamics of mRNA unwinding. Here, we present a comprehensive review of these models and also discuss other possible models. We evaluate each model and discuss the consistency and/or inconsistency between the theoretical results that are obtained based on each model and the available experimental data, thus determining which model is the most reasonable one to describe the mRNA unwinding mechanism and dynamics of the ribosome. Moreover, a framework for future studies in this subject is provided.
DOI
Optical Binding of Nanowires
Stephen H. Simpson, Pavel Zemánek, Onofrio M. Maragò, Philip H. Jones, and Simon Hanna
Multiple scattering of light induces structured interactions, or optical binding forces, between collections of small particles. This has been extensively studied in the case of microspheres. However, binding forces are strongly shape dependent: here, we turn our attention to dielectric nanowires. Using a novel numerical model we uncover rich behavior. The extreme geometry of the nanowires produces a sequence of stationary and dynamic states. In linearly polarized light, thermally stable ladder-like structures emerge. Lower symmetry, sagittate arrangements can also arise, whose configurational asymmetry unbalances the optical forces leading to nonconservative, translational motion. Finally, the addition of circular polarization drives a variety of coordinated rotational states whose dynamics expose fundamental properties of optical spin. These results suggest that optical binding can provide an increased level of control over the positions and motions of nanoparticles, opening new possibilities for driven self-organization and heralding a new field of self-assembling optically driven micromachines.
DOI
Multiple scattering of light induces structured interactions, or optical binding forces, between collections of small particles. This has been extensively studied in the case of microspheres. However, binding forces are strongly shape dependent: here, we turn our attention to dielectric nanowires. Using a novel numerical model we uncover rich behavior. The extreme geometry of the nanowires produces a sequence of stationary and dynamic states. In linearly polarized light, thermally stable ladder-like structures emerge. Lower symmetry, sagittate arrangements can also arise, whose configurational asymmetry unbalances the optical forces leading to nonconservative, translational motion. Finally, the addition of circular polarization drives a variety of coordinated rotational states whose dynamics expose fundamental properties of optical spin. These results suggest that optical binding can provide an increased level of control over the positions and motions of nanoparticles, opening new possibilities for driven self-organization and heralding a new field of self-assembling optically driven micromachines.
DOI
Nanomechanics of the substrate binding domain of Hsp70 determine its allosteric ATP-induced conformational change
Soumit Sankar Mandal, Dale R. Merz, Maximilian Buchsteiner, Ruxandra I. Dima, Matthias Rief, and Gabriel Žoldák
Owing to the cooperativity of protein structures, it is often almost impossible to identify independent subunits, flexible regions, or hinges simply by visual inspection of static snapshots. Here, we use single-molecule force experiments and simulations to apply tension across the substrate binding domain (SBD) of heat shock protein 70 (Hsp70) to pinpoint mechanical units and flexible hinges. The SBD consists of two nanomechanical units matching 3D structural parts, called the α- and β-subdomain. We identified a flexible region within the rigid β-subdomain that gives way under load, thus opening up the α/β interface. In exactly this region, structural changes occur in the ATP-induced opening of Hsp70 to allow substrate exchange. Our results show that the SBD’s ability to undergo large conformational changes is already encoded by passive mechanics of the individual elements.
DOI
Owing to the cooperativity of protein structures, it is often almost impossible to identify independent subunits, flexible regions, or hinges simply by visual inspection of static snapshots. Here, we use single-molecule force experiments and simulations to apply tension across the substrate binding domain (SBD) of heat shock protein 70 (Hsp70) to pinpoint mechanical units and flexible hinges. The SBD consists of two nanomechanical units matching 3D structural parts, called the α- and β-subdomain. We identified a flexible region within the rigid β-subdomain that gives way under load, thus opening up the α/β interface. In exactly this region, structural changes occur in the ATP-induced opening of Hsp70 to allow substrate exchange. Our results show that the SBD’s ability to undergo large conformational changes is already encoded by passive mechanics of the individual elements.
DOI
Brownian dynamics simulations to explore experimental microsphere diffusion with optical tweezers
Manuel Pancorbo, Miguel A. Rubio, P. Domínguez-García
We develop two-dimensional Brownian dynamics simulations to examine the motion of disks under thermal fluctuations and Hookean forces. Our simulations are designed to be experimental-like, since the experimental conditions define the available time-scales which characterize the solution of Langevin equations. To define the fluid model and methodology, we explain the basics of the theory of Brownian motion applicable to quasi-twodimensional diffusion of optically-trapped microspheres. Using the data produced by the simulations, we propose an alternative methodology to calculate diffusion coefficients. We obtain that, using typical input parameters in video-microscopy experiments, the averaged values of the diffusion coefficient differ from the theoretical one less than a 1%.
DOI
We develop two-dimensional Brownian dynamics simulations to examine the motion of disks under thermal fluctuations and Hookean forces. Our simulations are designed to be experimental-like, since the experimental conditions define the available time-scales which characterize the solution of Langevin equations. To define the fluid model and methodology, we explain the basics of the theory of Brownian motion applicable to quasi-twodimensional diffusion of optically-trapped microspheres. Using the data produced by the simulations, we propose an alternative methodology to calculate diffusion coefficients. We obtain that, using typical input parameters in video-microscopy experiments, the averaged values of the diffusion coefficient differ from the theoretical one less than a 1%.
DOI
Monday, June 19, 2017
Tomographic active optical trapping of arbitrarily shaped objects by exploiting 3D refractive index maps
Kyoohyun Kim & YongKeun Park
Optical trapping can manipulate the three-dimensional (3D) motion of spherical particles based on the simple prediction of optical forces and the responding motion of samples. However, controlling the 3D behaviour of non-spherical particles with arbitrary orientations is extremely challenging, due to experimental difficulties and extensive computations. Here, we achieve the real-time optical control of arbitrarily shaped particles by combining the wavefront shaping of a trapping beam and measurements of the 3D refractive index distribution of samples. Engineering the 3D light field distribution of a trapping beam based on the measured 3D refractive index map of samples generates a light mould, which can manipulate colloidal and biological samples with arbitrary orientations and/or shapes. The present method provides stable control of the orientation and assembly of arbitrarily shaped particles without knowing a priori information about the sample geometry. The proposed method can be directly applied in biophotonics and soft matter physics.
DOI
Optical trapping can manipulate the three-dimensional (3D) motion of spherical particles based on the simple prediction of optical forces and the responding motion of samples. However, controlling the 3D behaviour of non-spherical particles with arbitrary orientations is extremely challenging, due to experimental difficulties and extensive computations. Here, we achieve the real-time optical control of arbitrarily shaped particles by combining the wavefront shaping of a trapping beam and measurements of the 3D refractive index distribution of samples. Engineering the 3D light field distribution of a trapping beam based on the measured 3D refractive index map of samples generates a light mould, which can manipulate colloidal and biological samples with arbitrary orientations and/or shapes. The present method provides stable control of the orientation and assembly of arbitrarily shaped particles without knowing a priori information about the sample geometry. The proposed method can be directly applied in biophotonics and soft matter physics.
DOI
Versatile Gap Mode Plasmon under ATR Geometry towards Single Molecule Raman, Laser Trapping and Photocatalytic Reactions
Masayuki FUTAMATA, Keitaro AKAI, Chiaki IIDA, Natsumi AKIBA
We have investigated various aspects of a gap mode plasmon to establish it as an analytical tool. First, markedly large (107 – 109) enhancement factors for the Raman scattering intensity from a thiophenol (TP) monolayer sandwiched by Ag films on a prism and silver nanoparticles (AgNPs) were obtained under attenuated total reflection (ATR) geometry. Second, AgNPs with a radius of ∼20 nm were optically trapped and immobilized on TP-covered Ag films under a gap mode resonance with extremely weak laser power density of ∼1 μW/μm2 at 532 nm. The observed optical trapping and immobilization were theoretically rationalized using a dipole-dipole coupling and van der Waals interaction between AgNPs and Ag films. Third, p-alkyl TP molecules such as p-methyl TP, p-ethyl TP, p-isopropyl TP, and p-tertiary butyl TP were photocatalytically oxidized into p-carboxyl TP, whereas o- and m-methyl TP did not show such reactions.
DOI
We have investigated various aspects of a gap mode plasmon to establish it as an analytical tool. First, markedly large (107 – 109) enhancement factors for the Raman scattering intensity from a thiophenol (TP) monolayer sandwiched by Ag films on a prism and silver nanoparticles (AgNPs) were obtained under attenuated total reflection (ATR) geometry. Second, AgNPs with a radius of ∼20 nm were optically trapped and immobilized on TP-covered Ag films under a gap mode resonance with extremely weak laser power density of ∼1 μW/μm2 at 532 nm. The observed optical trapping and immobilization were theoretically rationalized using a dipole-dipole coupling and van der Waals interaction between AgNPs and Ag films. Third, p-alkyl TP molecules such as p-methyl TP, p-ethyl TP, p-isopropyl TP, and p-tertiary butyl TP were photocatalytically oxidized into p-carboxyl TP, whereas o- and m-methyl TP did not show such reactions.
DOI
Dual-trap system to study charged graphene nanoplatelets in high vacuum
Joyce E. Coppock, Pavel Nagornykh, Jacob P. J. Murphy, I. S. McAdams, Saimouli Katragadda, and B. E. Kane
We discuss the design and implementation of a system to generate charged multilayer graphene nanoplatelets and introduce a nanoplatelet into a quadrupole ion trap under vacuum. Levitation decouples the platelet from the environment and enables sensitive mechanical and magnetic measurements. The platelets are generated via liquid exfoliation of graphite pellets and charged via electrospray ionization. A single platelet is trapped at a pressure of several hundred millitorr and transferred to a trap in a second chamber, which is pumped to ultra high vacuum pressures for further study.
DOI
We discuss the design and implementation of a system to generate charged multilayer graphene nanoplatelets and introduce a nanoplatelet into a quadrupole ion trap under vacuum. Levitation decouples the platelet from the environment and enables sensitive mechanical and magnetic measurements. The platelets are generated via liquid exfoliation of graphite pellets and charged via electrospray ionization. A single platelet is trapped at a pressure of several hundred millitorr and transferred to a trap in a second chamber, which is pumped to ultra high vacuum pressures for further study.
DOI
Coherent control of a single nitrogen-vacancy center spin in optically levitated nanodiamond
Robert M. Pettit, Levi P. Neukirch, Yi Zhang, and A. Nick Vamivakas
We report the first observation, to the best of our knowledge, of electron spin transients in single negatively charged nitrogen-vacancy (NV−NV−) centers, contained within optically trapped nanodiamonds, in both atmospheric pressure and low vacuum. It is shown that, after an initial exposure to low vacuum, the trapped nanodiamonds remain at temperatures near room temperature even in low vacuum. Furthermore, the transverse coherence time of the NV−NV− center spin, measured to be 𝑇2=101.4 nsT2=101.4 ns, is robust over the range of trapping powers considered in this study.
DOI
We report the first observation, to the best of our knowledge, of electron spin transients in single negatively charged nitrogen-vacancy (NV−NV−) centers, contained within optically trapped nanodiamonds, in both atmospheric pressure and low vacuum. It is shown that, after an initial exposure to low vacuum, the trapped nanodiamonds remain at temperatures near room temperature even in low vacuum. Furthermore, the transverse coherence time of the NV−NV− center spin, measured to be 𝑇2=101.4 nsT2=101.4 ns, is robust over the range of trapping powers considered in this study.
DOI
Radiation pressure on a diffractive sailcraft
Grover A. Swartzlander
Advanced diffractive metamaterial films may afford advantages over passive reflective surfaces for a variety space missions that use solar or laser in-space propulsion. Three cases are compared: sun-facing diffractive sails, Littrow diffraction configurations, and conventional reflective sails. A simple Earth-to-Mars orbit transfer at a constant attitude with respect to the sunline finds no penalty for transparent diffractive sails. Advantages of the latter approach include actively controlled metasails, reuse of photons, and mission-specific optimization schemes.
DOI
Advanced diffractive metamaterial films may afford advantages over passive reflective surfaces for a variety space missions that use solar or laser in-space propulsion. Three cases are compared: sun-facing diffractive sails, Littrow diffraction configurations, and conventional reflective sails. A simple Earth-to-Mars orbit transfer at a constant attitude with respect to the sunline finds no penalty for transparent diffractive sails. Advantages of the latter approach include actively controlled metasails, reuse of photons, and mission-specific optimization schemes.
DOI
Wednesday, June 14, 2017
Coupling librational and translational motion of a levitated nanoparticle in an optical cavity
Shengyan Liu, Tongcang Li, and Zhang-qi Yin
An optically levitated nonspherical nanoparticle can exhibit both librational and translational vibrations due to orientational and translational confinements of the optical tweezer, respectively. Usually, the frequency of its librational mode in a linearly polarized optical tweezer is much larger than the frequency of its translational mode. Because of the frequency mismatch, the intrinsic coupling between librational and translational modes is very weak in vacuum. Here we propose a scheme to couple its librational and center-of-mass modes with an optical cavity mode. By adiabatically eliminating the cavity mode, the beam splitter Hamiltonian between librational and center-of-mass modes can be realized. We find that high-fidelity quantum state transfer between the librational and translational modes can be achieved with practical parameters. Our work may find applications in sympathetic cooling of multiple modes and quantum information processing.
DOI
An optically levitated nonspherical nanoparticle can exhibit both librational and translational vibrations due to orientational and translational confinements of the optical tweezer, respectively. Usually, the frequency of its librational mode in a linearly polarized optical tweezer is much larger than the frequency of its translational mode. Because of the frequency mismatch, the intrinsic coupling between librational and translational modes is very weak in vacuum. Here we propose a scheme to couple its librational and center-of-mass modes with an optical cavity mode. By adiabatically eliminating the cavity mode, the beam splitter Hamiltonian between librational and center-of-mass modes can be realized. We find that high-fidelity quantum state transfer between the librational and translational modes can be achieved with practical parameters. Our work may find applications in sympathetic cooling of multiple modes and quantum information processing.
DOI
Collapse-induced orientational localization of rigid rotors
Björn Schrinski, Benjamin A. Stickler, and Klaus Hornberger
We show how the ro-translational motion of anisotropic particles is affected by the model of continuous spontaneous localization (CSL), the most prominent hypothetical modification of the Schrödinger equation restoring realism on the macroscale. We derive the master equation describing collapse-induced spatio-orientational decoherence and demonstrate how it leads to linear- and angular-momentum diffusion. Since the associated heating rates scale differently with the CSL parameters, the latter can be determined individually by measuring the random motion of a single levitated nanorotor.
DOI
We show how the ro-translational motion of anisotropic particles is affected by the model of continuous spontaneous localization (CSL), the most prominent hypothetical modification of the Schrödinger equation restoring realism on the macroscale. We derive the master equation describing collapse-induced spatio-orientational decoherence and demonstrate how it leads to linear- and angular-momentum diffusion. Since the associated heating rates scale differently with the CSL parameters, the latter can be determined individually by measuring the random motion of a single levitated nanorotor.
DOI
Acoustic force measurements on polymer-coated microbubbles in a microfluidic device
Gianluca Memoli, Christopher R. Fury, Kate O. Baxter, and Pierre N. Gélat, Philip H. Jones
This work presents an acoustofluidic device for manipulating coated microbubbles, designed for the simultaneous use of optical and acoustical tweezers. A comprehensive characterization of the acoustic pressure in the device is presented, obtained by the synergic use of different techniques in the range of acoustic frequencies where visual observations showed aggregation of polymer-coated microbubbles. In absence of bubbles, the combined use of laser vibrometry and finite element modelling supported a non-invasive measurement of the acoustic pressure and an enhanced understanding of the system resonances. Calibrated holographic optical tweezers were used for direct measurements of the acoustic forces acting on an isolated microbubble, at low driving pressures, and to confirm the spatial distribution of the acoustic field. This allowed quantitative acoustic pressure measurements by particle tracking, using polystyrene beads, and an evaluation of the related uncertainties. This process facilitated the extension of tracking to microbubbles, which have a negative acoustophoretic contrast factor, allowing acoustic force measurements on bubbles at higher pressures than optical tweezers, highlighting four peaks in the acoustic response of the device. Results and methodologies are relevant to acoustofluidic applications requiring a precise characterization of the acoustic field and, in general, to biomedical applications with microbubbles or deformable particles.
DOI
This work presents an acoustofluidic device for manipulating coated microbubbles, designed for the simultaneous use of optical and acoustical tweezers. A comprehensive characterization of the acoustic pressure in the device is presented, obtained by the synergic use of different techniques in the range of acoustic frequencies where visual observations showed aggregation of polymer-coated microbubbles. In absence of bubbles, the combined use of laser vibrometry and finite element modelling supported a non-invasive measurement of the acoustic pressure and an enhanced understanding of the system resonances. Calibrated holographic optical tweezers were used for direct measurements of the acoustic forces acting on an isolated microbubble, at low driving pressures, and to confirm the spatial distribution of the acoustic field. This allowed quantitative acoustic pressure measurements by particle tracking, using polystyrene beads, and an evaluation of the related uncertainties. This process facilitated the extension of tracking to microbubbles, which have a negative acoustophoretic contrast factor, allowing acoustic force measurements on bubbles at higher pressures than optical tweezers, highlighting four peaks in the acoustic response of the device. Results and methodologies are relevant to acoustofluidic applications requiring a precise characterization of the acoustic field and, in general, to biomedical applications with microbubbles or deformable particles.
DOI
Conformational dynamics of the frameshift stimulatory structure in HIV-1
Dustin B Ritchie, Tonia R Cappellano, Collin Tittle, Negar Rezajooei, Logan Rouleau, William KA Sikkema and Michael T Woodside
Programmed ribosomal frameshifting (PRF) in HIV-1 is thought to be stimulated by a hairpin in the mRNA, although a pseudoknot-like triplex has also been proposed. Because the conformational dynamics of the stimulatory structure under tension applied by the ribosomal helicase during translation may play an important role in PRF, we used optical tweezers to apply tension to the HIV stimulatory structure and monitor its unfolding and refolding dynamics. The folding and unfolding kinetics and energy landscape of the hairpin were measured by ramping the force on the hairpin up and down, providing a detailed biophysical characterisation. Unexpectedly, whereas unfolding reflected the simple two-state behavior typical of many hairpins, refolding was more complex, displaying significant heterogeneity. Evidence was found for multiple refolding pathways as well as previously unsuspected, partially folded intermediates. Measuring a variant mRNA containing only the sequence required to form the proposed triplex, it behaved largely in the same way. Nonetheless, very rarely, high-force unfolding events characteristic of pseudoknot-like structures were observed. The rare occurrence of the triplex suggests that the hairpin is the functional stimulatory structure. The unusual heterogeneity of the hairpin dynamics under tension suggests a possible functional role in PRF similar to the dynamics of other stimulatory structures.
DOI
Programmed ribosomal frameshifting (PRF) in HIV-1 is thought to be stimulated by a hairpin in the mRNA, although a pseudoknot-like triplex has also been proposed. Because the conformational dynamics of the stimulatory structure under tension applied by the ribosomal helicase during translation may play an important role in PRF, we used optical tweezers to apply tension to the HIV stimulatory structure and monitor its unfolding and refolding dynamics. The folding and unfolding kinetics and energy landscape of the hairpin were measured by ramping the force on the hairpin up and down, providing a detailed biophysical characterisation. Unexpectedly, whereas unfolding reflected the simple two-state behavior typical of many hairpins, refolding was more complex, displaying significant heterogeneity. Evidence was found for multiple refolding pathways as well as previously unsuspected, partially folded intermediates. Measuring a variant mRNA containing only the sequence required to form the proposed triplex, it behaved largely in the same way. Nonetheless, very rarely, high-force unfolding events characteristic of pseudoknot-like structures were observed. The rare occurrence of the triplex suggests that the hairpin is the functional stimulatory structure. The unusual heterogeneity of the hairpin dynamics under tension suggests a possible functional role in PRF similar to the dynamics of other stimulatory structures.
DOI
Microfluidic-based high-throughput optical trapping of nanoparticles
Abhay Kotnala, Yi Zheng, Jianping Fu and Wei Cheng
Optical tweezers have emerged as a powerful tool for multiparametric analysis of individual nanoparticles with single-molecule sensitivity. However, its inherent low-throughput characteristic remains a major obstacle to its applications within and beyond the laboratory. This limitation is further exacerbated when working with low concentration nanoparticle samples. Here, we present a microfluidic-based optical tweezers system that can ‘actively’ deliver nanoparticles to a designated microfluidic region for optical trapping and analysis. The active microfluidic delivery of nanoparticles results in significantly improved throughput and efficiency for optical trapping of nanoparticles. We observed a more than tenfold increase in optical trapping throughput for nanoparticles as compared to conventional systems at the same nanoparticle concentration. To demonstrate the utility of this microfluidic-based optical tweezers system, we further used back-focal plane interferometry coupled with a trapping laser for the precise quantitation of nanoparticle size without prior knowledge of the refractive index of nanoparticles. The development of this microfluidic-based active optical tweezers system thus opens the door to high-throughput multiparametric analysis of nanoparticles using precision optical traps in the future.
DOI
Optical tweezers have emerged as a powerful tool for multiparametric analysis of individual nanoparticles with single-molecule sensitivity. However, its inherent low-throughput characteristic remains a major obstacle to its applications within and beyond the laboratory. This limitation is further exacerbated when working with low concentration nanoparticle samples. Here, we present a microfluidic-based optical tweezers system that can ‘actively’ deliver nanoparticles to a designated microfluidic region for optical trapping and analysis. The active microfluidic delivery of nanoparticles results in significantly improved throughput and efficiency for optical trapping of nanoparticles. We observed a more than tenfold increase in optical trapping throughput for nanoparticles as compared to conventional systems at the same nanoparticle concentration. To demonstrate the utility of this microfluidic-based optical tweezers system, we further used back-focal plane interferometry coupled with a trapping laser for the precise quantitation of nanoparticle size without prior knowledge of the refractive index of nanoparticles. The development of this microfluidic-based active optical tweezers system thus opens the door to high-throughput multiparametric analysis of nanoparticles using precision optical traps in the future.
DOI
Tuesday, June 13, 2017
Optical pulling and pushing forces exerted on silicon nanospheres with strong coherent interaction between electric and magnetic resonances
Hongfeng Liu, Mingcheng Panmai, Yuanyuan Peng, and Sheng Lan
We investigated theoretically and numerically the optical pulling and pushing forces acting on silicon (Si) nanospheres (NSs) with strong coherent interaction between electric and magnetic resonances. We examined the optical pulling and pushing forces exerted on Si NSs by two interfering waves and revealed the underlying physical mechanism from the viewpoint of electric- and magnetic-dipole manipulation. As compared with a polystyrene (PS) NS, it was found that the optical pulling force for a Si NS with the same size is enlarged by nearly two orders of magnitude. In addition to the optical pulling force appearing at the long-wavelength side of the magnetic dipole resonance, very large optical pushing force is observed at the magnetic quadrupole resonance. The correlation between the optical pulling/pushing force and the directional scattering characterized by the ratio of the forward to backward scattering was revealed. More interestingly, it was found that the high-order electric and magnetic resonances in large Si NSs play an important role in producing optical pulling force which can be generated by not only s-polarized wave but also p-polarized one. Our finding indicates that the strong coherent interaction between the electric and magnetic resonances existing in nanoparticles with large refractive indices can be exploited to manipulate the optical force acting on them and the correlation between the optical force and the directional scattering can be used as guidance. The engineering and manipulation of optical forces will find potential applications in the trapping, transport and sorting of nanoparticles.
DOI
We investigated theoretically and numerically the optical pulling and pushing forces acting on silicon (Si) nanospheres (NSs) with strong coherent interaction between electric and magnetic resonances. We examined the optical pulling and pushing forces exerted on Si NSs by two interfering waves and revealed the underlying physical mechanism from the viewpoint of electric- and magnetic-dipole manipulation. As compared with a polystyrene (PS) NS, it was found that the optical pulling force for a Si NS with the same size is enlarged by nearly two orders of magnitude. In addition to the optical pulling force appearing at the long-wavelength side of the magnetic dipole resonance, very large optical pushing force is observed at the magnetic quadrupole resonance. The correlation between the optical pulling/pushing force and the directional scattering characterized by the ratio of the forward to backward scattering was revealed. More interestingly, it was found that the high-order electric and magnetic resonances in large Si NSs play an important role in producing optical pulling force which can be generated by not only s-polarized wave but also p-polarized one. Our finding indicates that the strong coherent interaction between the electric and magnetic resonances existing in nanoparticles with large refractive indices can be exploited to manipulate the optical force acting on them and the correlation between the optical force and the directional scattering can be used as guidance. The engineering and manipulation of optical forces will find potential applications in the trapping, transport and sorting of nanoparticles.
DOI
Translation and folding of single proteins in real time
Florian Wruck, Alexandros Katranidis, Knud H. Nierhaus, Georg Büldt, and Martin Hegner
Protein biosynthesis is inherently coupled to cotranslational protein folding. Folding of the nascent chain already occurs during synthesis and is mediated by spatial constraints imposed by the ribosomal exit tunnel as well as self-interactions. The polypeptide’s vectorial emergence from the ribosomal tunnel establishes the possible folding pathways leading to its native tertiary structure. How cotranslational protein folding and the rate of synthesis are linked to a protein’s amino acid sequence is still not well defined. Here, we follow synthesis by individual ribosomes using dual-trap optical tweezers and observe simultaneous folding of the nascent polypeptide chain in real time. We show that observed stalling during translation correlates with slowed peptide bond formation at successive proline sequence positions and electrostatic interactions between positively charged amino acids and the ribosomal tunnel. We also determine possible cotranslational folding sites initiated by hydrophobic collapse for an unstructured and two globular proteins while directly measuring initial cotranslational folding forces. Our study elucidates the intricate relationship among a protein’s amino acid sequence, its cotranslational nascent-chain elongation rate, and folding.
DOI
Protein biosynthesis is inherently coupled to cotranslational protein folding. Folding of the nascent chain already occurs during synthesis and is mediated by spatial constraints imposed by the ribosomal exit tunnel as well as self-interactions. The polypeptide’s vectorial emergence from the ribosomal tunnel establishes the possible folding pathways leading to its native tertiary structure. How cotranslational protein folding and the rate of synthesis are linked to a protein’s amino acid sequence is still not well defined. Here, we follow synthesis by individual ribosomes using dual-trap optical tweezers and observe simultaneous folding of the nascent polypeptide chain in real time. We show that observed stalling during translation correlates with slowed peptide bond formation at successive proline sequence positions and electrostatic interactions between positively charged amino acids and the ribosomal tunnel. We also determine possible cotranslational folding sites initiated by hydrophobic collapse for an unstructured and two globular proteins while directly measuring initial cotranslational folding forces. Our study elucidates the intricate relationship among a protein’s amino acid sequence, its cotranslational nascent-chain elongation rate, and folding.
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Adaptive and Specific Recognition of Telomeric G-Quadruplexes via Polyvalency Induced Unstacking of Binding Units
Jibin Abraham Punnoose , Yue Ma, Yuanyuan Li, Mai Sakuma, Shankar Mandal, Kazuo Nagasawa, and Hanbin Mao
Targeting DNA G-quadruplexes using small-molecule ligands has shown to modulate biological functions mediated by G-quadruplexes inside cells. Given >716 000 G-quadruplex hosting sites in human genome, the specific binding of ligands to quadruplex becomes problematic. Here, we innovated a polyvalency based mechanism to specifically target multiple telomeric G-quadruplexes. We synthesized a tetrameric telomestatin derivative and evaluated its complex polyvalent binding with multiple G-quadruplexes by single-molecule mechanical unfolding in laser tweezers. We found telomestatin tetramer binds to multimeric telomeric G-quadruplexes >40 times stronger than monomeric quadruplexes, which can be ascribed to the polyvalency induced unstacking of binding units (or PIU binding) for G-quadruplexes. While stacking of telomestatin units in the tetramer imparts steric hindrance for the ligand to access stand-alone G-quadruplexes, the stacking disassembles to accommodate the potent polyvalent binding between the tetramer ligand and multimeric G-quadruplexes. We anticipate this adaptive PIU binding offers a generic mechanism to selectively target polymeric biomolecules prevalent inside cells.
DOI
Targeting DNA G-quadruplexes using small-molecule ligands has shown to modulate biological functions mediated by G-quadruplexes inside cells. Given >716 000 G-quadruplex hosting sites in human genome, the specific binding of ligands to quadruplex becomes problematic. Here, we innovated a polyvalency based mechanism to specifically target multiple telomeric G-quadruplexes. We synthesized a tetrameric telomestatin derivative and evaluated its complex polyvalent binding with multiple G-quadruplexes by single-molecule mechanical unfolding in laser tweezers. We found telomestatin tetramer binds to multimeric telomeric G-quadruplexes >40 times stronger than monomeric quadruplexes, which can be ascribed to the polyvalency induced unstacking of binding units (or PIU binding) for G-quadruplexes. While stacking of telomestatin units in the tetramer imparts steric hindrance for the ligand to access stand-alone G-quadruplexes, the stacking disassembles to accommodate the potent polyvalent binding between the tetramer ligand and multimeric G-quadruplexes. We anticipate this adaptive PIU binding offers a generic mechanism to selectively target polymeric biomolecules prevalent inside cells.
DOI
Ripping RNA by Force Using Gaussian Network Models
Changbong Hyeon and D. Thirumala
Using force as a probe to map the folding landscapes of RNA molecules has become a reality thanks to major advances in single molecule pulling experiments. Although the unfolding pathways under tension are complicated to predict, studies in the context of proteins have shown that topology is the major determinant of the unfolding landscapes. By building on this finding we study the responses of RNA molecules to force by adapting Gaussian network model (GNM) that represents RNAs using a bead–spring network with isotropic interactions. Cross-correlation matrices of residue fluctuations, which are analytically calculated using GNM even upon application of mechanical force, show distinct allosteric communication as RNAs rupture. The model is used to calculate the force–extension curves at full thermodynamic equilibrium, and the corresponding unfolding pathways of four RNA molecules subject to a quasi-statically increased force. Our study finds that the analysis using GNM captures qualitatively the unfolding pathway of T. ribozyme elucidated by the optical tweezers measurement. However, the simple model cannot capture features, such as bifurcation in the unfolding pathways or the ion effects, in the forced-unfolding of RNAs.
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Direct observation of coupling between orientation and flow fluctuations in a nematic liquid crystal at equilibrium
Hiroshi Orihara, Nobutaka Sakurai, Yuji Sasaki, and Tomoyuki Nagaya
To demonstrate coupling between orientation and flow fluctuations in a nematic liquid crystal at equilibrium, we simultaneously observe the intensity change due to director fluctuations under a polarizing microscope and the Brownian motion of a fluorescent particle trapped weakly by optical tweezers. The calculated cross-correlation function of the particle position and the spatial gradient of the intensity is nonzero, clearly indicating the existence of coupling.
DOI
To demonstrate coupling between orientation and flow fluctuations in a nematic liquid crystal at equilibrium, we simultaneously observe the intensity change due to director fluctuations under a polarizing microscope and the Brownian motion of a fluorescent particle trapped weakly by optical tweezers. The calculated cross-correlation function of the particle position and the spatial gradient of the intensity is nonzero, clearly indicating the existence of coupling.
DOI
Surface plasmon resonance in gold nanoparticles: a review
Vincenzo Amendola, Roberto Pilot, Marco Frasconi, Onofrio M Maragò and Maria Antonia Iatì
In the last two decades, plasmon resonance in gold nanoparticles (Au NPs) has been the subject of intense research efforts. Plasmon physics is intriguing and its precise modelling proved to be challenging. In fact, plasmons are highly responsive to a multitude of factors, either intrinsic to the Au NPs or from the environment, and recently the need emerged for the correction of standard electromagnetic approaches with quantum effects. Applications related to plasmon absorption and scattering in Au NPs are impressively numerous, ranging from sensing to photothermal effects to cell imaging. Also, plasmon-enhanced phenomena are highly interesting for multiple purposes, including, for instance, Raman spectroscopy of nearby analytes, catalysis, or sunlight energy conversion. In addition, plasmon excitation is involved in a series of advanced physical processes such as non-linear optics, optical trapping, magneto-plasmonics, and optical activity. Here, we provide the general overview of the field and the background for appropriate modelling of the physical phenomena. Then, we report on the current state of the art and most recent applications of plasmon resonance in Au NPs.
DOI
In the last two decades, plasmon resonance in gold nanoparticles (Au NPs) has been the subject of intense research efforts. Plasmon physics is intriguing and its precise modelling proved to be challenging. In fact, plasmons are highly responsive to a multitude of factors, either intrinsic to the Au NPs or from the environment, and recently the need emerged for the correction of standard electromagnetic approaches with quantum effects. Applications related to plasmon absorption and scattering in Au NPs are impressively numerous, ranging from sensing to photothermal effects to cell imaging. Also, plasmon-enhanced phenomena are highly interesting for multiple purposes, including, for instance, Raman spectroscopy of nearby analytes, catalysis, or sunlight energy conversion. In addition, plasmon excitation is involved in a series of advanced physical processes such as non-linear optics, optical trapping, magneto-plasmonics, and optical activity. Here, we provide the general overview of the field and the background for appropriate modelling of the physical phenomena. Then, we report on the current state of the art and most recent applications of plasmon resonance in Au NPs.
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Dual-Mode Fiber Optofluidic Flowmeter With a Large Dynamic Range
Yuan Gong ; Liming Qiu ; Chenlin Zhang ; Yu Wu ; Yun-Jiang Rao ; Gang-Ding Peng
A dual-mode fiber optofluidic flowmeter with a large dynamic range of four orders of magnitude is developed. The sensing mechanism is based on the force balance on an optically trapped polystyrene microparticle. As the optical force is at piconewton level, the flowmeter is very sensitive with a lower detection limit of 10 nl/min. In the open-loop mode, the manipulation length is used as the sensing output and the flowmeter has an inverse sensitivity so that it has higher sensitivity at lower flow rate. In the closed-loop mode, the manipulation length is set constant and a feedback signal, tuning the laser power for the force balance, is used as the sensing output. The closed-loop mode is helpful for extending the upper detection limit of flow rate and also enhancing the sensitivity at higher flow rate. This paper introduces a new kind of high-performance optofluidic sensors based on optical forces.
DOI
A dual-mode fiber optofluidic flowmeter with a large dynamic range of four orders of magnitude is developed. The sensing mechanism is based on the force balance on an optically trapped polystyrene microparticle. As the optical force is at piconewton level, the flowmeter is very sensitive with a lower detection limit of 10 nl/min. In the open-loop mode, the manipulation length is used as the sensing output and the flowmeter has an inverse sensitivity so that it has higher sensitivity at lower flow rate. In the closed-loop mode, the manipulation length is set constant and a feedback signal, tuning the laser power for the force balance, is used as the sensing output. The closed-loop mode is helpful for extending the upper detection limit of flow rate and also enhancing the sensitivity at higher flow rate. This paper introduces a new kind of high-performance optofluidic sensors based on optical forces.
DOI
Monday, June 12, 2017
DNA synthesis determines the binding mode of the human mitochondrial single-stranded DNA-binding protein
José A. Morin Fernando Cerrón Javier Jarillo Elena Beltran-Heredia Grzegorz L. Ciesielski J. Ricardo Arias-Gonzalez Laurie S. Kaguni Francisco J. Cao Borja Ibarra
Single-stranded DNA-binding proteins (SSBs) play a key role in genome maintenance, binding and organizing single-stranded DNA (ssDNA) intermediates. Multimeric SSBs, such as the human mitochondrial SSB (HmtSSB), present multiple sites to interact with ssDNA, which has been shown in vitro to enable them to bind a variable number of single-stranded nucleotides depending on the salt and protein concentration. It has long been suggested that different binding modes might be used selectively for different functions. To study this possibility, we used optical tweezers to determine and compare the structure and energetics of long, individual HmtSSB–DNA complexes assembled on preformed ssDNA and on ssDNA generated gradually during ‘in situ’ DNA synthesis. We show that HmtSSB binds to preformed ssDNA in two major modes, depending on salt and protein concentration. However, when protein binding was coupled to strand-displacement DNA synthesis, only one of the two binding modes was observed under all experimental conditions. Our results reveal a key role for the gradual generation of ssDNA in modulating the binding mode of a multimeric SSB protein and consequently, in generating the appropriate nucleoprotein structure for DNA synthetic reactions required for genome maintenance.
DOI
Single-stranded DNA-binding proteins (SSBs) play a key role in genome maintenance, binding and organizing single-stranded DNA (ssDNA) intermediates. Multimeric SSBs, such as the human mitochondrial SSB (HmtSSB), present multiple sites to interact with ssDNA, which has been shown in vitro to enable them to bind a variable number of single-stranded nucleotides depending on the salt and protein concentration. It has long been suggested that different binding modes might be used selectively for different functions. To study this possibility, we used optical tweezers to determine and compare the structure and energetics of long, individual HmtSSB–DNA complexes assembled on preformed ssDNA and on ssDNA generated gradually during ‘in situ’ DNA synthesis. We show that HmtSSB binds to preformed ssDNA in two major modes, depending on salt and protein concentration. However, when protein binding was coupled to strand-displacement DNA synthesis, only one of the two binding modes was observed under all experimental conditions. Our results reveal a key role for the gradual generation of ssDNA in modulating the binding mode of a multimeric SSB protein and consequently, in generating the appropriate nucleoprotein structure for DNA synthetic reactions required for genome maintenance.
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DNA origami supported precision measurements of biomolecular interactions and structure
Hendrik Dietz
Programmable self-assembly with DNA origami allows creating custom-shaped nanoscale objects. Through this capacity, DNA origami enables constructing custom instruments to perform precision measurements of molecular interactions and structure, with enhanced control over positioning, orientating and manipulating the molecules under study. In my presentation I will report about a series of experiments in which we exploited this capacity to dissect the weak stacking forces between individual basepairs (1) and the forces that act between pairs of nucleosomes (2).
In experiment (1), we directly measured at the single-molecule level the forces and lifetimes of DNA base-pair stacking interactions for all stack sequence combinations. Our experimental approach combined dual-beam optical tweezers with DNA origami components to allow positioning of blunt-end DNA helices so that the weak stacking force could be isolated. Base-pair stack arrays that lacked a covalent backbone connection spontaneously dissociated at average rates ranging from 0.02 to 500 per second, depending on the sequence combination and stack array size. Forces in the range from 2 to 8 piconewtons that act along the helical direction only mildly accelerated the stochastic unstacking process. The free-energy increments per stack that we estimate from the measured forward and backward kinetic rates ranged from −0.8 to −3.4 kilocalories per mole, depending on the sequence combination. Our data contributes to understanding the mechanics of DNA processing in biology, and it is helpful for designing the kinetics of DNA-based nanoscale devices according to user specifications.
In experiment (2), we performed a direct measurement of inter-nucleosomal interactions by integrating two nucleosomes into a DNA origami-based force spectrometer, which enabled sub-nanometer resolution measurements of nucleosome-nucleosome distance frequencies via single particle electron microscopy imaging. From the data we derived the Boltzmann-weighted distance-dependent energy landscape for nucleosome pair interactions. We find a shallow but long-ranged (~6 nm) attractive nucleosome pair potential with a minimum of −1.6 kcal/mol close to direct-contact distances. The relative nucleosome orientation had little influence, but histone H4 acetylation or removal of histone tails drastically decreased the interaction strength. Due to the weak and shallow pair potential, higher-order nucleosome assemblies will be compliant and experience dynamic shape fluctuations in the absence of additional co-factors. Our results contribute to a more accurate description of chromatin, while our force spectrometer provides a powerful tool for the direct and high-resolution study of molecular interactions using imaging techniques.
DOI
Programmable self-assembly with DNA origami allows creating custom-shaped nanoscale objects. Through this capacity, DNA origami enables constructing custom instruments to perform precision measurements of molecular interactions and structure, with enhanced control over positioning, orientating and manipulating the molecules under study. In my presentation I will report about a series of experiments in which we exploited this capacity to dissect the weak stacking forces between individual basepairs (1) and the forces that act between pairs of nucleosomes (2).
In experiment (1), we directly measured at the single-molecule level the forces and lifetimes of DNA base-pair stacking interactions for all stack sequence combinations. Our experimental approach combined dual-beam optical tweezers with DNA origami components to allow positioning of blunt-end DNA helices so that the weak stacking force could be isolated. Base-pair stack arrays that lacked a covalent backbone connection spontaneously dissociated at average rates ranging from 0.02 to 500 per second, depending on the sequence combination and stack array size. Forces in the range from 2 to 8 piconewtons that act along the helical direction only mildly accelerated the stochastic unstacking process. The free-energy increments per stack that we estimate from the measured forward and backward kinetic rates ranged from −0.8 to −3.4 kilocalories per mole, depending on the sequence combination. Our data contributes to understanding the mechanics of DNA processing in biology, and it is helpful for designing the kinetics of DNA-based nanoscale devices according to user specifications.
In experiment (2), we performed a direct measurement of inter-nucleosomal interactions by integrating two nucleosomes into a DNA origami-based force spectrometer, which enabled sub-nanometer resolution measurements of nucleosome-nucleosome distance frequencies via single particle electron microscopy imaging. From the data we derived the Boltzmann-weighted distance-dependent energy landscape for nucleosome pair interactions. We find a shallow but long-ranged (~6 nm) attractive nucleosome pair potential with a minimum of −1.6 kcal/mol close to direct-contact distances. The relative nucleosome orientation had little influence, but histone H4 acetylation or removal of histone tails drastically decreased the interaction strength. Due to the weak and shallow pair potential, higher-order nucleosome assemblies will be compliant and experience dynamic shape fluctuations in the absence of additional co-factors. Our results contribute to a more accurate description of chromatin, while our force spectrometer provides a powerful tool for the direct and high-resolution study of molecular interactions using imaging techniques.
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Mechanical untying of the smallest knotted protein from different pulling axis using optical tweezers
Maira Rivera, Andrés Bustamante, Yuxin Hao, Rodrigo A Maillard and Mauricio Baez
Knotted polypeptides constitute a group of proteins whose backbone chain entangles in the folded state. The folding mechanism of knotted protein is complex, involving multiple intermediate states. Therefore, it is difficult to determine experimentally the thermodynamic and kinetic barriers associated with threading the polypeptide chain. To address this problem, we used optical tweezers to study the folding mechanism of MJ0366, the smallest protein containing a shallow trefoil knot. Specifically, we mechanically manipulated the knotted protein from different pulling directions to either tighten the knot upon unfolding or to untie the protein from different points of its structure. When the knot was tightened in force-ramp experiments, we observed single unfolding and refolding transitions characterized by unimodal force distributions, and an accompanying change in contour length (Lc = 24 ± 3 nm) consistent with the expected theoretical value. Moreover, in constant-force experiments, we observed that the protein oscillates between two states, whose frequency of interconversion showed a linear dependence with force. These observations suggest a two-state folding mechanism when the trefoil knot remains in the polypeptide chain. However, when the knot is untied by keeping the C-termini free to thread, the protein unfolds in either one or two transitions and at forces higher than those observed when pulling from the N and C termini. Moreover, 50% of the unfolding transitions have a Lc shorter than the expected value, suggesting that the protein can be trapped in a misfolded state during its refolding process. When the knot is untied by keeping both N- and C-termini free to thread, we observed a single unfolding force distribution with the expected Lc for this construct. However, the refolding process could not be characterized because it occurs at forces below the resolution limit (<1 pN). Our results suggest that when the knot remains in the unfolded state, the folding landscape have a smooth and funneled shape. However, when the knotted protein has to overcome the threading process this energy landscape becomes rough.
DOI
Knotted polypeptides constitute a group of proteins whose backbone chain entangles in the folded state. The folding mechanism of knotted protein is complex, involving multiple intermediate states. Therefore, it is difficult to determine experimentally the thermodynamic and kinetic barriers associated with threading the polypeptide chain. To address this problem, we used optical tweezers to study the folding mechanism of MJ0366, the smallest protein containing a shallow trefoil knot. Specifically, we mechanically manipulated the knotted protein from different pulling directions to either tighten the knot upon unfolding or to untie the protein from different points of its structure. When the knot was tightened in force-ramp experiments, we observed single unfolding and refolding transitions characterized by unimodal force distributions, and an accompanying change in contour length (Lc = 24 ± 3 nm) consistent with the expected theoretical value. Moreover, in constant-force experiments, we observed that the protein oscillates between two states, whose frequency of interconversion showed a linear dependence with force. These observations suggest a two-state folding mechanism when the trefoil knot remains in the polypeptide chain. However, when the knot is untied by keeping the C-termini free to thread, the protein unfolds in either one or two transitions and at forces higher than those observed when pulling from the N and C termini. Moreover, 50% of the unfolding transitions have a Lc shorter than the expected value, suggesting that the protein can be trapped in a misfolded state during its refolding process. When the knot is untied by keeping both N- and C-termini free to thread, we observed a single unfolding force distribution with the expected Lc for this construct. However, the refolding process could not be characterized because it occurs at forces below the resolution limit (<1 pN). Our results suggest that when the knot remains in the unfolded state, the folding landscape have a smooth and funneled shape. However, when the knotted protein has to overcome the threading process this energy landscape becomes rough.
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The Folding Mechanism of an Artificial Knotted Protein Characterized by Optical Tweezers
Andrés Bustamante, Maira Rivera, José Molina and Mauricio Baez
Protein folding occurs because establishment of weak interactions observed in the native state (native contacts) predominate over the formation of transient interactions that could promote alternative conformations (non-native contacts). However, knotted proteins defy this view because their polypeptide chains would require to form non-native contacts during its folding. To determine the role of non-native contacts in such topologies, we explored the folding mechanism of an artificial knotted protein (2ouf-knot). In this case, optimization of non-native contacts should play a minor role during its folding. The folding of 2ouf-knot was characterized maintaining the knotted topology in the unfolded state by pulling their C and N extremes by using optical tweezers. The molecular stretching of 2ouf-knot obtained at constant velocity of pulling showed two transitions of unfolding occurring at 9 ± 1 and 14 ± 1 pN, followed by two transitions of refolding at 8 ± 1 and 12 ± 1 pN. This indicates the presence of an intermediate in both directions of the folding reaction of 2ouf-knot. In terms of the molecular extension, the contour length (Lc) associated with the first transition at low force account for the 30% of the expected extension for the fully unfolded polypeptide. Notably, the intermediate of 2ouf-knot was also apparent under equilibrium conditions. At constant force, 2ouf-knot oscillates between the intermediate and the unfolded state extended by 5 and 16 nm with respect to the native state respectively. We observe that there was no full unfolding of the protein without visiting the intermediate state of 5 nm. Therefore, this specie is an obligate step (on-pathway intermediate) for the complete unfolding of the protein. Nevertheless, we also observe that approximately 20% of the times the fully unfolded protein jump until reach a smaller intermediate of 4 nm with very short residence time. This short-live off-pathway intermediates backtrack to the fully unfolded state which refolds to the on-pathway intermediate of 5 nm. These results contrast with the absence of intermediates (on and off-pathway) in the case of a natural knotted protein (MJ0366 of Methanocaldococcus jannaschii) characterized under the same experimental setup. We suggest that the intermediates observed in the case of artificial knots arise by lack of specific non-native contacts that could help to accommodate the knot during unfolding/refolding process. These specific interactions otherwise will help to smooth the energetic landscape of natural knotted proteins.
DOI
Protein folding occurs because establishment of weak interactions observed in the native state (native contacts) predominate over the formation of transient interactions that could promote alternative conformations (non-native contacts). However, knotted proteins defy this view because their polypeptide chains would require to form non-native contacts during its folding. To determine the role of non-native contacts in such topologies, we explored the folding mechanism of an artificial knotted protein (2ouf-knot). In this case, optimization of non-native contacts should play a minor role during its folding. The folding of 2ouf-knot was characterized maintaining the knotted topology in the unfolded state by pulling their C and N extremes by using optical tweezers. The molecular stretching of 2ouf-knot obtained at constant velocity of pulling showed two transitions of unfolding occurring at 9 ± 1 and 14 ± 1 pN, followed by two transitions of refolding at 8 ± 1 and 12 ± 1 pN. This indicates the presence of an intermediate in both directions of the folding reaction of 2ouf-knot. In terms of the molecular extension, the contour length (Lc) associated with the first transition at low force account for the 30% of the expected extension for the fully unfolded polypeptide. Notably, the intermediate of 2ouf-knot was also apparent under equilibrium conditions. At constant force, 2ouf-knot oscillates between the intermediate and the unfolded state extended by 5 and 16 nm with respect to the native state respectively. We observe that there was no full unfolding of the protein without visiting the intermediate state of 5 nm. Therefore, this specie is an obligate step (on-pathway intermediate) for the complete unfolding of the protein. Nevertheless, we also observe that approximately 20% of the times the fully unfolded protein jump until reach a smaller intermediate of 4 nm with very short residence time. This short-live off-pathway intermediates backtrack to the fully unfolded state which refolds to the on-pathway intermediate of 5 nm. These results contrast with the absence of intermediates (on and off-pathway) in the case of a natural knotted protein (MJ0366 of Methanocaldococcus jannaschii) characterized under the same experimental setup. We suggest that the intermediates observed in the case of artificial knots arise by lack of specific non-native contacts that could help to accommodate the knot during unfolding/refolding process. These specific interactions otherwise will help to smooth the energetic landscape of natural knotted proteins.
DOI
Inter-Domain Interactions In Nascent Polypeptides Interfere With Productive Protein Folding
Kaixian Liu, Kevin Maciuba and Christian Kaiser
How large, multi-domain proteins robustly fold into their native structures is not well understood. It is often assumed that individual domains fold independently, but experimental studies have largely been limited to the special case of tandem repeat proteins. We have used single-molecule force spectroscopy to study the folding of elongation factor G (EF-G), which is composed of five domains. Optical tweezers experiments show that full-length EF-G refolds inefficiently along highly complex folding pathways. To determine whether vectorial protein synthesis, by reducing complexity, enables efficient domain-wise folding, we studied successively longer ribosome-bound nascent polypeptides. The N-terminal G-domain forms a stable structure by itself and enables the otherwise unstable subsequent domain II to fold. When both domains fold simultaneously, they form a misfolded species that is more stable than the productive folding intermediates. Interestingly, interactions with the unfolded domain II destabilize the native G-domain. Together, our data indicate that vectorial protein synthesis, perhaps with rates optimized for folding, helps to avoid intra-molecular misfolding within nascent multi-domain polypeptides, allowing the G-domain to complete folding before domain II is produced. Interactions with molecular chaperones might be required to protect folded domains against interactions with yet unstructured regions in the nascent polypeptide.
DOI
How large, multi-domain proteins robustly fold into their native structures is not well understood. It is often assumed that individual domains fold independently, but experimental studies have largely been limited to the special case of tandem repeat proteins. We have used single-molecule force spectroscopy to study the folding of elongation factor G (EF-G), which is composed of five domains. Optical tweezers experiments show that full-length EF-G refolds inefficiently along highly complex folding pathways. To determine whether vectorial protein synthesis, by reducing complexity, enables efficient domain-wise folding, we studied successively longer ribosome-bound nascent polypeptides. The N-terminal G-domain forms a stable structure by itself and enables the otherwise unstable subsequent domain II to fold. When both domains fold simultaneously, they form a misfolded species that is more stable than the productive folding intermediates. Interestingly, interactions with the unfolded domain II destabilize the native G-domain. Together, our data indicate that vectorial protein synthesis, perhaps with rates optimized for folding, helps to avoid intra-molecular misfolding within nascent multi-domain polypeptides, allowing the G-domain to complete folding before domain II is produced. Interactions with molecular chaperones might be required to protect folded domains against interactions with yet unstructured regions in the nascent polypeptide.
DOI
Dual function of the trigger factor chaperone in nascent protein folding
Christian Kaiser, Kevin Maciuba and Kaixian Liu
Multi-domain proteins often require help from molecular chaperones to fold productively, even before the ribosome has finished their synthesis. The mechanisms underlying chaperone function remain poorly understood. Using optical tweezers to study the folding of elongation factor G (EF-G), a model multi-domain protein, we find that the N-terminal G-domain in nascent EF-G polypeptides folds robustly. The following domain II, in contrast, fails to fold efficiently. Strikingly, interactions with the unfolded domain II convert the natively folded G domain to a non-native state. This non-native state readily unfolds, and the two unfolded domains subsequently form misfolded states, preventing productive folding. Both the conversion of natively folded domains and non-productive interactions among unfolded domains are efficiently prevented by the nascent chain-binding chaperone trigger factor. Thus, our single-molecule measurements of multi-domain protein folding reveal an unexpected role for the chaperone: It protects already folded domains against denaturation resulting from interactions with parts of the nascent polypeptide that are not folded yet. Previous studies had implicated trigger factor in guiding the folding of individual domains, and interactions among domains had been neglected. Avoiding these early folding defects is crucial, since they can propagate and result in misfolding of the entire protein.
DOI
Multi-domain proteins often require help from molecular chaperones to fold productively, even before the ribosome has finished their synthesis. The mechanisms underlying chaperone function remain poorly understood. Using optical tweezers to study the folding of elongation factor G (EF-G), a model multi-domain protein, we find that the N-terminal G-domain in nascent EF-G polypeptides folds robustly. The following domain II, in contrast, fails to fold efficiently. Strikingly, interactions with the unfolded domain II convert the natively folded G domain to a non-native state. This non-native state readily unfolds, and the two unfolded domains subsequently form misfolded states, preventing productive folding. Both the conversion of natively folded domains and non-productive interactions among unfolded domains are efficiently prevented by the nascent chain-binding chaperone trigger factor. Thus, our single-molecule measurements of multi-domain protein folding reveal an unexpected role for the chaperone: It protects already folded domains against denaturation resulting from interactions with parts of the nascent polypeptide that are not folded yet. Previous studies had implicated trigger factor in guiding the folding of individual domains, and interactions among domains had been neglected. Avoiding these early folding defects is crucial, since they can propagate and result in misfolding of the entire protein.
DOI
Friday, June 9, 2017
Thermally induced micro-motion by inflection in optical potential
Martin Šiler, Petr Jákl, Oto Brzobohatý, Artem Ryabov, Radim Filip & Pavel Zemánek
Recent technological progress in a precise control of optically trapped objects allows much broader ventures to unexplored territory of thermal motion in non-linear potentials. In this work, we exploit an experimental set-up of holographic optical tweezers to experimentally investigate Brownian motion of a micro-particle near the inflection point of the cubic optical potential. We present two complementary views on the non-linear Brownian motion. On an ensemble of stochastic trajectories, we simultaneously determine (i) the detailed short-time position statistics and (ii) the long-distance first-passage time statistics. We evaluate specific statistical moment ratios demonstrating strongly non-linear stochastic dynamics. This is a crucial step towards a possible massive exploitation of the broad class of complex non-linear stochastic effects with objects of more complex structure and shape including living ones.
DOI
Recent technological progress in a precise control of optically trapped objects allows much broader ventures to unexplored territory of thermal motion in non-linear potentials. In this work, we exploit an experimental set-up of holographic optical tweezers to experimentally investigate Brownian motion of a micro-particle near the inflection point of the cubic optical potential. We present two complementary views on the non-linear Brownian motion. On an ensemble of stochastic trajectories, we simultaneously determine (i) the detailed short-time position statistics and (ii) the long-distance first-passage time statistics. We evaluate specific statistical moment ratios demonstrating strongly non-linear stochastic dynamics. This is a crucial step towards a possible massive exploitation of the broad class of complex non-linear stochastic effects with objects of more complex structure and shape including living ones.
DOI
Optomechanical measurement of the role of lamins in whole cell deformability
Thorsten Kolb, Julia Kraxner, Kai Skodzek, Michael Haug, Dean Crawford, Kendra K. Maaß, Katerina E. Aifantis, Graeme Whyte
There is mounting evidence that the nuclear envelope, and particularly the lamina, plays a critical role in the mechanical and regulation properties of the cell and changes to the lamina can have implications for the physical properties of the whole cell. In this study we demonstrate that the optical stretcher can measure changes in the time-dependent mechanical properties of living cells with different levels of A-type lamin expression. Results from the optical stretcher shows a decrease in the deformability of cells as the levels of lamin A increases, for cells which grow both adherently and in suspension. Further detail can be probed by combining the optical stretcher with fluorescence microscopy to investigate the nuclear mechanical properties which show a larger decrease in deformability than for the whole cell.
DOI
There is mounting evidence that the nuclear envelope, and particularly the lamina, plays a critical role in the mechanical and regulation properties of the cell and changes to the lamina can have implications for the physical properties of the whole cell. In this study we demonstrate that the optical stretcher can measure changes in the time-dependent mechanical properties of living cells with different levels of A-type lamin expression. Results from the optical stretcher shows a decrease in the deformability of cells as the levels of lamin A increases, for cells which grow both adherently and in suspension. Further detail can be probed by combining the optical stretcher with fluorescence microscopy to investigate the nuclear mechanical properties which show a larger decrease in deformability than for the whole cell.
DOI
A Single Spherical Assembly of Protein Amyloid Fibrils Formed by Laser Trapping
Dr. Ken-ichi Yuyama, Mariko Ueda, Dr. Satoshi Nagao, Prof. Shun Hirota, Prof. Teruki Sugiyama, Prof. Hiroshi Masuhara
Protein amyloids have received much attention owing to their correlation with serious diseases and to their promising mechanical and optical properties as future materials. Amyloid formation has been conducted by tuning temperature and chemical conditions, so that its nucleation and the following growth are analyzed as ensemble dynamics. A single spherical assembly of amyloid fibrils of cytochrome c domain-swapped dimer was successfully generated upon laser trapping. The amyloid fibrillar structure was confirmed by fluorescence characterization and electron microscopy. The prepared spheres were further manipulated individually in solution to fabricate a three-dimensional microstructure and a line pattern. Amyloid formation dynamics and amyloid-based microstructure fabrication are demonstrated based on direct observation of a single spherical assembly, which foresees a new approach in amyloid studies.
DOI
Protein amyloids have received much attention owing to their correlation with serious diseases and to their promising mechanical and optical properties as future materials. Amyloid formation has been conducted by tuning temperature and chemical conditions, so that its nucleation and the following growth are analyzed as ensemble dynamics. A single spherical assembly of amyloid fibrils of cytochrome c domain-swapped dimer was successfully generated upon laser trapping. The amyloid fibrillar structure was confirmed by fluorescence characterization and electron microscopy. The prepared spheres were further manipulated individually in solution to fabricate a three-dimensional microstructure and a line pattern. Amyloid formation dynamics and amyloid-based microstructure fabrication are demonstrated based on direct observation of a single spherical assembly, which foresees a new approach in amyloid studies.
DOI
Membrane curvature regulates ligand-specific membrane sorting of GPCRs in living cells
adla R Rosholm, Natascha Leijnse, Anna Mantsiou, Vadym Tkach, Søren L Pedersen, Volker F Wirth, Lene B Oddershede, Knud J Jensen, Karen L Martinez, Nikos S Hatzakis, Poul Martin Bendix, Andrew Callan-Jones & Dimitrios Stamou
The targeted spatial organization (sorting) of Gprotein-coupled receptors (GPCRs) is essential for their biological function and often takes place in highly curved membrane compartments such as filopodia, endocytic pits, trafficking vesicles or endosome tubules. However, the influence of geometrical membrane curvature on GPCR sorting remains unknown. Here we used fluorescence imaging to establish a quantitative correlation between membrane curvature and sorting of three prototypic class A GPCRs (the neuropeptide Y receptor Y2, the β1 adrenergic receptor and the β2 adrenergic receptor) in living cells. Fitting of a thermodynamic model to the data enabled us to quantify how sorting is mediated by an energetic drive to match receptor shape and membrane curvature. Curvature-dependent sorting was regulated by ligands in a specific manner. We anticipate that this curvature-dependent biomechanical coupling mechanism contributes to the sorting, trafficking and function of transmembrane proteins in general.
DOI
The targeted spatial organization (sorting) of Gprotein-coupled receptors (GPCRs) is essential for their biological function and often takes place in highly curved membrane compartments such as filopodia, endocytic pits, trafficking vesicles or endosome tubules. However, the influence of geometrical membrane curvature on GPCR sorting remains unknown. Here we used fluorescence imaging to establish a quantitative correlation between membrane curvature and sorting of three prototypic class A GPCRs (the neuropeptide Y receptor Y2, the β1 adrenergic receptor and the β2 adrenergic receptor) in living cells. Fitting of a thermodynamic model to the data enabled us to quantify how sorting is mediated by an energetic drive to match receptor shape and membrane curvature. Curvature-dependent sorting was regulated by ligands in a specific manner. We anticipate that this curvature-dependent biomechanical coupling mechanism contributes to the sorting, trafficking and function of transmembrane proteins in general.
DOI
Optical levitation of a mirror for reaching the standard quantum limit
Yuta Michimura, Yuya Kuwahara, Takafumi Ushiba, Nobuyuki Matsumoto, and Masaki Ando
We propose a new method to optically levitate a macroscopic mirror with two vertical Fabry-Pérot cavities linearly aligned. This configuration gives the simplest possible optical levitation in which the number of laser beams used is the minimum of two. We demonstrate that reaching the standard quantum limit (SQL) of a displacement measurement with our system is feasible with current technology. The cavity geometry and the levitated mirror parameters are designed to ensure that the Brownian vibration of the mirror surface is smaller than the SQL. Our scheme provides a promising tool for testing macroscopic quantum mechanics.
DOI
DOI
Wednesday, June 7, 2017
Effective Light Directed Assembly of Building Blocks with Microscale Control
Ngoc-Duy Dinh, Rongcong Luo, Maria Tankeh Asuncion Christine, Weikang Nicholas Lin, Wei-Chuan Shih, James Cho-Hong Goh, Chia-Hung Chen
Light-directed forces have been widely used to pattern micro/nanoscale objects with precise control, forming functional assemblies. However, a substantial laser intensity is required to generate sufficient optical gradient forces to move a small object in a certain direction, causing limited throughput for applications. A high-throughput light-directed assembly is demonstrated as a printing technology by introducing gold nanorods to induce thermal convection flows that move microparticles (diameter = 40 µm to several hundreds of micrometers) to specific light-guided locations, forming desired patterns. With the advantage of effective light-directed assembly, the microfluidic-fabricated monodispersed biocompatible microparticles are used as building blocks to construct a structured assembly (≈10 cm scale) in ≈2 min. The control with microscale precision is approached by changing the size of the laser light spot. After crosslinking assembly of building blocks, a novel soft material with wanted pattern is approached. To demonstrate its application, the mesenchymal stem-cell-seeded hydrogel microparticles are prepared as functional building blocks to construct scaffold-free tissues with desired structures. This light-directed fabrication method can be applied to integrate different building units, enabling the bottom-up formation of materials with precise control over their internal structure for bioprinting, tissue engineering, and advanced manufacturing.
DOI
Light-directed forces have been widely used to pattern micro/nanoscale objects with precise control, forming functional assemblies. However, a substantial laser intensity is required to generate sufficient optical gradient forces to move a small object in a certain direction, causing limited throughput for applications. A high-throughput light-directed assembly is demonstrated as a printing technology by introducing gold nanorods to induce thermal convection flows that move microparticles (diameter = 40 µm to several hundreds of micrometers) to specific light-guided locations, forming desired patterns. With the advantage of effective light-directed assembly, the microfluidic-fabricated monodispersed biocompatible microparticles are used as building blocks to construct a structured assembly (≈10 cm scale) in ≈2 min. The control with microscale precision is approached by changing the size of the laser light spot. After crosslinking assembly of building blocks, a novel soft material with wanted pattern is approached. To demonstrate its application, the mesenchymal stem-cell-seeded hydrogel microparticles are prepared as functional building blocks to construct scaffold-free tissues with desired structures. This light-directed fabrication method can be applied to integrate different building units, enabling the bottom-up formation of materials with precise control over their internal structure for bioprinting, tissue engineering, and advanced manufacturing.
DOI
Experimental observation of Shapiro-steps in colloidal monolayers driven across time-dependent substrate potentials
T. Brazda, C. July and C. Bechinger
We experimentally study the motion of a colloidal monolayer which is driven across a commensurate substrate potential whose amplitude is periodically modulated in time. In addition to a significant reduction of the static friction force compared to an unmodulated substrate, we observe a Shapiro step structure in the force dependence of the mean particle velocity which is explained by the dynamical mode locking between the particle motion and the substrate modulation. In this regime, the entire crystal moves in a stick-slip fashion similar to what is observed when a single point contact is driven across a periodic surface. Contrary to numerical simulations, where typically a large number of Shapiro steps is found, only a single step is observed in our experiments. This is explained by the formation of kinks which weaken the synchronization between adjacent particles.
DOI
We experimentally study the motion of a colloidal monolayer which is driven across a commensurate substrate potential whose amplitude is periodically modulated in time. In addition to a significant reduction of the static friction force compared to an unmodulated substrate, we observe a Shapiro step structure in the force dependence of the mean particle velocity which is explained by the dynamical mode locking between the particle motion and the substrate modulation. In this regime, the entire crystal moves in a stick-slip fashion similar to what is observed when a single point contact is driven across a periodic surface. Contrary to numerical simulations, where typically a large number of Shapiro steps is found, only a single step is observed in our experiments. This is explained by the formation of kinks which weaken the synchronization between adjacent particles.
DOI
Radiation force and torque of light-sheets
F G Mitri
The aim of this work is to provide exact analytical closed-form expressions for the longitudinal and transverse optical radiation force and axial spin torque components, for a 2D surface cross-section with arbitrary shape in the field of light-sheet beams of arbitrary wavefront. Generalized partial-wave series expressions for the longitudinal and transverse optical radiation forces and torque are derived based on the multipole expansion in cylindrical wave functions, stemming from series expansions for the incident and scattered electromagnetic fields. The incident light-sheet wavefields are expressed using generalized series involving the beam-shape coefficients (BSCs), and the scattered fields are given in terms of series involving the scattering coefficients of the object. Numerical illustrative examples on a dielectric absorptive circular cylindrical cross-section are provided for different wavefronts, ranging from plane waves, as well as non-paraxial scalar Airy and Gaussian light-sheet beams. The BSCs are derived based on the angular spectrum decomposition method, which provides adequate means to evaluate the radiation force and torque components when the cylinder cross-section is centered on the beam, or shifted off-axially with respect to the incident axis of wave propagation. In essence, the present theoretical analysis provides a complete formalism in the framework of the generalized Lorenz–Mie theory in 2D based upon exact closed-form series expressions to compute the optical force and torque components induced by 2D light-sheets of arbitrary wavefronts, illuminating a scatterer with an arbitrary geometrical cross-section (in 2D). Possible applications are in particle transport and rotation.
DOI
The aim of this work is to provide exact analytical closed-form expressions for the longitudinal and transverse optical radiation force and axial spin torque components, for a 2D surface cross-section with arbitrary shape in the field of light-sheet beams of arbitrary wavefront. Generalized partial-wave series expressions for the longitudinal and transverse optical radiation forces and torque are derived based on the multipole expansion in cylindrical wave functions, stemming from series expansions for the incident and scattered electromagnetic fields. The incident light-sheet wavefields are expressed using generalized series involving the beam-shape coefficients (BSCs), and the scattered fields are given in terms of series involving the scattering coefficients of the object. Numerical illustrative examples on a dielectric absorptive circular cylindrical cross-section are provided for different wavefronts, ranging from plane waves, as well as non-paraxial scalar Airy and Gaussian light-sheet beams. The BSCs are derived based on the angular spectrum decomposition method, which provides adequate means to evaluate the radiation force and torque components when the cylinder cross-section is centered on the beam, or shifted off-axially with respect to the incident axis of wave propagation. In essence, the present theoretical analysis provides a complete formalism in the framework of the generalized Lorenz–Mie theory in 2D based upon exact closed-form series expressions to compute the optical force and torque components induced by 2D light-sheets of arbitrary wavefronts, illuminating a scatterer with an arbitrary geometrical cross-section (in 2D). Possible applications are in particle transport and rotation.
DOI
Matter-Wave Tractor Beams
Alexey A. Gorlach, Maxim A. Gorlach, Andrei V. Lavrinenko, and Andrey Novitsky
Optical and acoustic tractor beams are currently the focus of intense research due to their counterintuitive property of exerting a pulling force on small scattering objects. In this Letter we propose a matter-wave tractor beam and utilize the de Broglie waves of nonrelativistic matter particles in analogy to “classical” tractor beams. We reveal the presence of the quantum-mechanical pulling force for the variety of quantum mechanical potentials observing the resonant enhancement of the pulling effect under the conditions of the suppressed scattering known as the Ramsauer-Townsend effect. We also derive the sufficient conditions on the scattering potential for the emergence of the pulling force and show that, in particular, a Coulomb scatterer is always shoved, while a Yukawa (screened Coulomb) scatterer can be drawn. Pulling forces in optics, acoustics, quantum mechanics, and classical mechanics are compared, and the matter-wave pulling force is found to have exclusive properties of dragging slow particles in short-range potentials. We envisage that the use of tractor beams could lead to the unprecedented precision in manipulation with atomic-scale quantum objects.
DOI
Optical and acoustic tractor beams are currently the focus of intense research due to their counterintuitive property of exerting a pulling force on small scattering objects. In this Letter we propose a matter-wave tractor beam and utilize the de Broglie waves of nonrelativistic matter particles in analogy to “classical” tractor beams. We reveal the presence of the quantum-mechanical pulling force for the variety of quantum mechanical potentials observing the resonant enhancement of the pulling effect under the conditions of the suppressed scattering known as the Ramsauer-Townsend effect. We also derive the sufficient conditions on the scattering potential for the emergence of the pulling force and show that, in particular, a Coulomb scatterer is always shoved, while a Yukawa (screened Coulomb) scatterer can be drawn. Pulling forces in optics, acoustics, quantum mechanics, and classical mechanics are compared, and the matter-wave pulling force is found to have exclusive properties of dragging slow particles in short-range potentials. We envisage that the use of tractor beams could lead to the unprecedented precision in manipulation with atomic-scale quantum objects.
DOI
The ribosome destabilizes native and non-native structures in a nascent multidomain protein
Kaixian Liu, Joseph E. Rehfus, Elliot Mattson, Christian M. Kaiser
Correct folding is a prerequisite for the biological activity of most proteins. Folding has largely been studied using in vitro refolding assays with isolated small, robustly folding proteins. A substantial fraction of all cellular proteomes is composed of multidomain proteins that are often not amenable to this approach, and their folding remains poorly understood. These large proteins likely begin to fold during their synthesis by the ribosome, a large molecular machine that translates the genetic code. The ribosome affects how folding proceeds, but the underlying mechanisms remain largely obscure. We have utilized optical tweezers to study the folding of elongation factor G, a multidomain protein composed of five domains. We find that interactions among unfolded domains interfere with productive folding in the full-length protein. The N-terminal G-domain constitutes an independently folding unit that, upon in vitro refolding, adopts two similar states that correspond to the natively folded and a non-native, possibly misfolded structure. The ribosome destabilizes both of these states, suggesting a mechanism by which terminal misfolding into highly stable, non-native structures is avoided. The ribosome may thus directly contribute to efficient folding by modulating the folding of nascent multidomain proteins.
DOI
Correct folding is a prerequisite for the biological activity of most proteins. Folding has largely been studied using in vitro refolding assays with isolated small, robustly folding proteins. A substantial fraction of all cellular proteomes is composed of multidomain proteins that are often not amenable to this approach, and their folding remains poorly understood. These large proteins likely begin to fold during their synthesis by the ribosome, a large molecular machine that translates the genetic code. The ribosome affects how folding proceeds, but the underlying mechanisms remain largely obscure. We have utilized optical tweezers to study the folding of elongation factor G, a multidomain protein composed of five domains. We find that interactions among unfolded domains interfere with productive folding in the full-length protein. The N-terminal G-domain constitutes an independently folding unit that, upon in vitro refolding, adopts two similar states that correspond to the natively folded and a non-native, possibly misfolded structure. The ribosome destabilizes both of these states, suggesting a mechanism by which terminal misfolding into highly stable, non-native structures is avoided. The ribosome may thus directly contribute to efficient folding by modulating the folding of nascent multidomain proteins.
DOI
Tuesday, June 6, 2017
Spatial distributions of pericellular stiffness in natural extracellular matrices are dependent on cell-mediated proteolysis and contractility
M. Keating, A. Kurup, M. Alvarez-Elizondo, A.J. Levine, E. Botvinick
Bulk tissue stiffness has been correlated with regulation of cellular processes and conversely cells have been shown to remodel their pericellular tissue according to a complex feedback mechanism critical to development, homeostasis, and disease. However, bulk rheological methods mask the dynamics within a heterogeneous fibrous extracellular matrix (ECM) in the region proximal to a cell (pericellular region). Here, we use optical tweezers active microrheology (AMR) to probe the distribution of the complex material response function (α = α′ + α″, in units of µm/nN) within a type I collagen ECM, a biomaterial commonly used in tissue engineering. We discovered cells both elastically and plastically deformed the pericellular material. α′ is wildly heterogeneous, with 1/α′ values spanning three orders of magnitude around a single cell. This was observed in gels having a cell-free 1/α′ of approximately 0.5 nN/µm. We also found that inhibition of cell contractility instantaneously softens the pericellular space and reduces stiffness heterogeneity, suggesting the system was strain hardened and not only plastically remodeled. The remaining regions of high stiffness suggest cellular remodeling of the surrounding matrix. To test this hypothesis, cells were incubated within the type I collagen gel for 24-h in a media containing a broad-spectrum matrix metalloproteinase (MMP) inhibitor. While pericellular material maintained stiffness asymmetry, stiffness magnitudes were reduced. Dual inhibition demonstrates that the combination of MMP activity and contractility is necessary to establish the pericellular stiffness landscape. This heterogeneity in stiffness suggests the distribution of pericellular stiffness, and not bulk stiffness alone, must be considered in the study of cell-ECM interactions and design of complex biomaterial scaffolds.
DOI
Bulk tissue stiffness has been correlated with regulation of cellular processes and conversely cells have been shown to remodel their pericellular tissue according to a complex feedback mechanism critical to development, homeostasis, and disease. However, bulk rheological methods mask the dynamics within a heterogeneous fibrous extracellular matrix (ECM) in the region proximal to a cell (pericellular region). Here, we use optical tweezers active microrheology (AMR) to probe the distribution of the complex material response function (α = α′ + α″, in units of µm/nN) within a type I collagen ECM, a biomaterial commonly used in tissue engineering. We discovered cells both elastically and plastically deformed the pericellular material. α′ is wildly heterogeneous, with 1/α′ values spanning three orders of magnitude around a single cell. This was observed in gels having a cell-free 1/α′ of approximately 0.5 nN/µm. We also found that inhibition of cell contractility instantaneously softens the pericellular space and reduces stiffness heterogeneity, suggesting the system was strain hardened and not only plastically remodeled. The remaining regions of high stiffness suggest cellular remodeling of the surrounding matrix. To test this hypothesis, cells were incubated within the type I collagen gel for 24-h in a media containing a broad-spectrum matrix metalloproteinase (MMP) inhibitor. While pericellular material maintained stiffness asymmetry, stiffness magnitudes were reduced. Dual inhibition demonstrates that the combination of MMP activity and contractility is necessary to establish the pericellular stiffness landscape. This heterogeneity in stiffness suggests the distribution of pericellular stiffness, and not bulk stiffness alone, must be considered in the study of cell-ECM interactions and design of complex biomaterial scaffolds.
DOI
Diagnosis of malarial infection using change in properties of optically trapped red blood cells
Apurba Paul, Ponnan Padmapriya, Vasant Natarajan
In previous work studying the properties of red blood cells (RBCs) held in an optical tweezers trap, we observed an increase in the spectrum of Brownian fluctuations for RBCs from a Plasmodium falciparum culture—due to increased rigidity of the cells—compared to normal RBCs. We wanted to extend the study to patient samples, since the earlier work was done with cultures grown in the lab. Individual RBCs were held in an optical-tweezers trap. Its position fluctuations were measured and the power spectrum determined. The corner frequency (fc) of the spectrum gave a quantitative measurement of the spectrum. The value of fc was 25 Hz for normal cells, which increased to 29 Hz for infected cells—both for P. falciparum and Plasmodium vivax infections. The technique of measuring fc can be used as a screening tool for malaria in patients with fever, since RBCs not carrying the parasite will also show the change due to the bystander effect, irrespective of whether it is caused by P. falciparum or P. vivax.
DOI
In previous work studying the properties of red blood cells (RBCs) held in an optical tweezers trap, we observed an increase in the spectrum of Brownian fluctuations for RBCs from a Plasmodium falciparum culture—due to increased rigidity of the cells—compared to normal RBCs. We wanted to extend the study to patient samples, since the earlier work was done with cultures grown in the lab. Individual RBCs were held in an optical-tweezers trap. Its position fluctuations were measured and the power spectrum determined. The corner frequency (fc) of the spectrum gave a quantitative measurement of the spectrum. The value of fc was 25 Hz for normal cells, which increased to 29 Hz for infected cells—both for P. falciparum and Plasmodium vivax infections. The technique of measuring fc can be used as a screening tool for malaria in patients with fever, since RBCs not carrying the parasite will also show the change due to the bystander effect, irrespective of whether it is caused by P. falciparum or P. vivax.
DOI
Theoretical investigation on optical Kerr effect in femtosecond laser trapping of dielectric microspheres
Anita Devi and Arijit K De
Stable trapping of individual dielectric nanoparticles under high-repetition-rate ultrafast pulsed excitation has been demonstrated and theoretically explained based on repetitive instantaneous trapping as well as optical nonlinearity assisted trapping. Here we estimate the force exerted on a micron-sized spherical dielectric particle including optical Kerr effect. Using geometric optics approximation, we show how inclusion of optical Kerr effect results in significant change of the force curves along axial direction under femtosecond pulsed excitation compared with continuous-wave excitation. The results show excellent agreement with previous experimental findings. Most importantly, similar to the optical trapping of nanoparticles under ultrafast pulsed excitation, we show that the efficiency of trapping of micron-sized particles is also governed by the barrier height (and not the absolute depth) of the axial trapping potential.
DOI
Stable trapping of individual dielectric nanoparticles under high-repetition-rate ultrafast pulsed excitation has been demonstrated and theoretically explained based on repetitive instantaneous trapping as well as optical nonlinearity assisted trapping. Here we estimate the force exerted on a micron-sized spherical dielectric particle including optical Kerr effect. Using geometric optics approximation, we show how inclusion of optical Kerr effect results in significant change of the force curves along axial direction under femtosecond pulsed excitation compared with continuous-wave excitation. The results show excellent agreement with previous experimental findings. Most importantly, similar to the optical trapping of nanoparticles under ultrafast pulsed excitation, we show that the efficiency of trapping of micron-sized particles is also governed by the barrier height (and not the absolute depth) of the axial trapping potential.
DOI
Plasmon-assisted optical trapping and anti-trapping
Aliaksandra Ivinskaya, Mihail I Petrov, Andrey A Bogdanov, Ivan Shishkin, Pavel Ginzburg and Alexander S Shalin
The ability to manipulate small objects with focused laser beams has opened a venue for investigating dynamical phenomena relevant to both fundamental and applied science. Nanophotonic and plasmonic structures enable superior performance in optical trapping via highly confined near-fields. In this case, the interplay between the excitation field, re-scattered fields and the eigenmodes of a structure can lead to remarkable effects; one such effect, as reported here, is particle trapping by laser light in a vicinity of metal surface. Surface plasmon excitation at the metal substrate plays a key role in tailoring the optical forces acting on a nearby particle. Depending on whether the illuminating Gaussian beam is focused above or below the metal-dielectric interface, an order-of-magnitude enhancement or reduction of the trap stiffness is achieved compared with that of standard glass substrates. Furthermore, a novel plasmon-assisted anti-trapping effect (particle repulsion from the beam axis) is predicted and studied. A highly accurate particle sorting scheme based on the new anti-trapping effect is analyzed. The ability to distinguish and configure various electromagnetic channels through the developed analytical theory provides guidelines for designing auxiliary nanostructures and achieving ultimate control over mechanical motion at the micro- and nano-scales.
DOI
The ability to manipulate small objects with focused laser beams has opened a venue for investigating dynamical phenomena relevant to both fundamental and applied science. Nanophotonic and plasmonic structures enable superior performance in optical trapping via highly confined near-fields. In this case, the interplay between the excitation field, re-scattered fields and the eigenmodes of a structure can lead to remarkable effects; one such effect, as reported here, is particle trapping by laser light in a vicinity of metal surface. Surface plasmon excitation at the metal substrate plays a key role in tailoring the optical forces acting on a nearby particle. Depending on whether the illuminating Gaussian beam is focused above or below the metal-dielectric interface, an order-of-magnitude enhancement or reduction of the trap stiffness is achieved compared with that of standard glass substrates. Furthermore, a novel plasmon-assisted anti-trapping effect (particle repulsion from the beam axis) is predicted and studied. A highly accurate particle sorting scheme based on the new anti-trapping effect is analyzed. The ability to distinguish and configure various electromagnetic channels through the developed analytical theory provides guidelines for designing auxiliary nanostructures and achieving ultimate control over mechanical motion at the micro- and nano-scales.
DOI
A liquid thermal gradient refractive index lens and using it to trap single living cell in flowing environments
H. L. Liu, Y. Shi, L. Liang, L. Li, S. S. Guo, L. Yin and Y. Yang
A gradient refractive index (GRIN) lens has a great potential for on-chip imaging and detection systems because of its flat surface with reduced defects. This paper reports a liquid thermal GRIN lens prepared using heat conduction between only one liquid, and uses it as a tunable optical tweezer for single living cell trapping in a flowing environment. This liquid GRIN lens consists of a trapezoidal region in the upper layer which is used to establish a GRIN profile by the heat conduction between three streams of benzyl alcohol with different temperatures, and subsequently a rhombus region in the lower layer with compensation liquids to form a steady square-law parabolic refractive index profile only in transverse direction. Simulations and experiments successfully show the real-time tunability of the focusing properties. The focal length can be modulated in the range of 500 μm with the minimum focal length of 430 μm. A considerable high enhancement factor achieves 5.4 whereas the full width at half maximum is 4 μm. The response time of the GRIN lens is about 20 ms. Based on this enhancement, tunable optical trapping for single human embryonic kidney 293 cell in the range of 280 μm is demonstrated by varying the focal length and working distance which is difficult for solid optical tweezers. The considerable quality of this liquid GRIN lens indicates on-chip applications especially in high quality optical imaging, detection and cells' handling.
DOI
A gradient refractive index (GRIN) lens has a great potential for on-chip imaging and detection systems because of its flat surface with reduced defects. This paper reports a liquid thermal GRIN lens prepared using heat conduction between only one liquid, and uses it as a tunable optical tweezer for single living cell trapping in a flowing environment. This liquid GRIN lens consists of a trapezoidal region in the upper layer which is used to establish a GRIN profile by the heat conduction between three streams of benzyl alcohol with different temperatures, and subsequently a rhombus region in the lower layer with compensation liquids to form a steady square-law parabolic refractive index profile only in transverse direction. Simulations and experiments successfully show the real-time tunability of the focusing properties. The focal length can be modulated in the range of 500 μm with the minimum focal length of 430 μm. A considerable high enhancement factor achieves 5.4 whereas the full width at half maximum is 4 μm. The response time of the GRIN lens is about 20 ms. Based on this enhancement, tunable optical trapping for single human embryonic kidney 293 cell in the range of 280 μm is demonstrated by varying the focal length and working distance which is difficult for solid optical tweezers. The considerable quality of this liquid GRIN lens indicates on-chip applications especially in high quality optical imaging, detection and cells' handling.
DOI
Optical stretching in continuous flows
Erik Wilfried Morawetz, Roland Stange, Tobias Kiessling, Jörg Schnauß and Josef Kaes
Rheology of living cells has developed an increasing need for high throughput measurements. Diseases such as cancer heavily remodel the cytoskeleton and impinge on cellular functions. Cells affected by such diseases show altered rheologic responses on many different levels rendering cells' mechanical fingerprints - a potential target for diagnostics. To counteract naturally occurring distributions of properties in samples of living cells and foster the validity of experiments, high numbers of single cell measurements are necessary. Here, we present the "in flow optical stretcher" (IFOS), a concept of non-invasive optical cytometry capable of high throughput rates, while working in a regime of long measurement times and low frequencies. The setup deforms whole cells in a continuous flow by optical forces, bypassing steps of cell positioning that are unavoidable in state-of-the-art optical stretcher devices. A prototype was built using polydimethylsiloxane soft lithography. In a proof of premise experiment, we show that in the IFOS it is possible to deform cells of mammalian origin which have been treated with cytochalasin. All recorded successful experiments took place in less than 2 s each, as opposed to 10-20 s in state-of-the-art optical stretcher devices. Although other microfluidic rheology devices achieve significantly higher throughput rates, they operate in different frequency regimes and probe different mechanical responses. The IFOS still captures viscoelastic properties and active responses of cells while aiming to maximize the throughput at creep times on the order of seconds. It can be assumed that an automatic IFOS reaches a throughput an order of magnitude higher than current devices that are based on optical stretching for cell rheology.
DOI
Rheology of living cells has developed an increasing need for high throughput measurements. Diseases such as cancer heavily remodel the cytoskeleton and impinge on cellular functions. Cells affected by such diseases show altered rheologic responses on many different levels rendering cells' mechanical fingerprints - a potential target for diagnostics. To counteract naturally occurring distributions of properties in samples of living cells and foster the validity of experiments, high numbers of single cell measurements are necessary. Here, we present the "in flow optical stretcher" (IFOS), a concept of non-invasive optical cytometry capable of high throughput rates, while working in a regime of long measurement times and low frequencies. The setup deforms whole cells in a continuous flow by optical forces, bypassing steps of cell positioning that are unavoidable in state-of-the-art optical stretcher devices. A prototype was built using polydimethylsiloxane soft lithography. In a proof of premise experiment, we show that in the IFOS it is possible to deform cells of mammalian origin which have been treated with cytochalasin. All recorded successful experiments took place in less than 2 s each, as opposed to 10-20 s in state-of-the-art optical stretcher devices. Although other microfluidic rheology devices achieve significantly higher throughput rates, they operate in different frequency regimes and probe different mechanical responses. The IFOS still captures viscoelastic properties and active responses of cells while aiming to maximize the throughput at creep times on the order of seconds. It can be assumed that an automatic IFOS reaches a throughput an order of magnitude higher than current devices that are based on optical stretching for cell rheology.
DOI
Monday, June 5, 2017
Wavelength-Dependent Plasmon-Mediated Coalescence of Two Gold Nanorods
Jiunn-Woei Liaw, Wu-Chun Lin & Mao-Kuen Kuo
Plasmon-mediated coalescence of two nearby gold nanorods (NRs) suspended in water induced by the illumination of a linearly polarized (LP) light was studied theoretically. We analyzed the coupled optical forces and torques in terms of Maxwell’s stress tensor upon two identical NRs irradiated by a LP plane wave using the multiple multipole method to estimate the optomechanical outcome. Numerical results show that the light-matter interaction can perform attraction or repulsion, depending on their initial configurations. For the attraction, the end-to-end or side-by-side coalescence of the two gold NRs could be caused by the LP light, depending on the wavelength. For example, the side-by-side coalescence of two adjacent NRs of r = 15 nm and L = 120 nm is most likely induced by 800-nm LP laser beam, whereas the end-to-end coalescence by 1064-nm or 1700-nm LP laser. These distinct phenomena are attributed to the perpendicular or parallel alignment of NR to the polarization of LP light in different wavelength ranges. The magnitude of optical force, proportional to the light’s fluence, could be stronger than van der Waals force. The estimation based on quasi-static model without considering the fluid dynamics may provide an insight to optical manipulation on the self-assembly of gold colloid.
DOI
Plasmon-mediated coalescence of two nearby gold nanorods (NRs) suspended in water induced by the illumination of a linearly polarized (LP) light was studied theoretically. We analyzed the coupled optical forces and torques in terms of Maxwell’s stress tensor upon two identical NRs irradiated by a LP plane wave using the multiple multipole method to estimate the optomechanical outcome. Numerical results show that the light-matter interaction can perform attraction or repulsion, depending on their initial configurations. For the attraction, the end-to-end or side-by-side coalescence of the two gold NRs could be caused by the LP light, depending on the wavelength. For example, the side-by-side coalescence of two adjacent NRs of r = 15 nm and L = 120 nm is most likely induced by 800-nm LP laser beam, whereas the end-to-end coalescence by 1064-nm or 1700-nm LP laser. These distinct phenomena are attributed to the perpendicular or parallel alignment of NR to the polarization of LP light in different wavelength ranges. The magnitude of optical force, proportional to the light’s fluence, could be stronger than van der Waals force. The estimation based on quasi-static model without considering the fluid dynamics may provide an insight to optical manipulation on the self-assembly of gold colloid.
DOI
Diversity of Knot Solitons in Liquid Crystals Manifested by Linking of Preimages in Torons and Hopfions
Paul J. Ackerman and Ivan I. Smalyukh
Topological solitons are knots in continuous physical fields classified by nonzero Hopf index values. Despite arising in theories that span many branches of physics, from elementary particles to condensed matter and cosmology, they remain experimentally elusive and poorly understood. We introduce a method of experimental and numerical analysis of such localized structures in liquid crystals that, similar to the mathematical Hopf maps, relates all points of the medium’s order parameter space to their closed-loop preimages within the three-dimensional solitons. We uncover a surprisingly large diversity of naturally occurring and laser-generated topologically nontrivial solitons with differently knotted nematic fields, which previously have not been realized in theories and experiments alike. We discuss the implications of the liquid crystal’s nonpolar nature on the knot soliton topology and how the medium’s chirality, confinement, and elastic anisotropy help to overcome the constraints of the Hobart-Derrick theorem, yielding static three-dimensional solitons without or with additional defects. Our findings will establish chiral nematics as a model system for experimental exploration of topological solitons and may impinge on understanding of such nonsingular field configurations in other branches of physics, as well as may lead to technological applications.
DOI
Topological solitons are knots in continuous physical fields classified by nonzero Hopf index values. Despite arising in theories that span many branches of physics, from elementary particles to condensed matter and cosmology, they remain experimentally elusive and poorly understood. We introduce a method of experimental and numerical analysis of such localized structures in liquid crystals that, similar to the mathematical Hopf maps, relates all points of the medium’s order parameter space to their closed-loop preimages within the three-dimensional solitons. We uncover a surprisingly large diversity of naturally occurring and laser-generated topologically nontrivial solitons with differently knotted nematic fields, which previously have not been realized in theories and experiments alike. We discuss the implications of the liquid crystal’s nonpolar nature on the knot soliton topology and how the medium’s chirality, confinement, and elastic anisotropy help to overcome the constraints of the Hobart-Derrick theorem, yielding static three-dimensional solitons without or with additional defects. Our findings will establish chiral nematics as a model system for experimental exploration of topological solitons and may impinge on understanding of such nonsingular field configurations in other branches of physics, as well as may lead to technological applications.
DOI
Dynamics Study of the Deformation of Red Blood Cell by Optical Tweezers
Konin E. Jean Michel, Yale Pavel, Megnassan Eugene, Michel A. Kouacou, Jeremie T. Zoueu
In recent years, extensive research has been carried out on red blood cells in order to investigate their mechanical properties. The interest in these studies has been possible thanks to the technological innovations made in the field of micro or nano manipulation of biological and non-biological particles without physical contact. In the present project, we have developed a new approach to study the deformation of red blood cells moving against a trapped microbead by applying a sinusoidal voltage (DC offset 3.5 Vpp) to the stage at 0.4 Hz frequencies. The oscillating movement imposed on the stage highlights the indentation test and the tensile test known for the study of mechanical behavior of materials. The mechanical properties found are: the modulus of elasticity (Young Modulus), the shear modulus, the coefficient of hardening and erythrocyte resistance coefficient. The axial shear modulus 25.00 ± 1.5 μN/m and the transversal shear modulus 15.7 ± 4.63 μN/m were compared to those in the literature. These values were respectively determined by Hooke’s law and the Hertz model.
DOI
In recent years, extensive research has been carried out on red blood cells in order to investigate their mechanical properties. The interest in these studies has been possible thanks to the technological innovations made in the field of micro or nano manipulation of biological and non-biological particles without physical contact. In the present project, we have developed a new approach to study the deformation of red blood cells moving against a trapped microbead by applying a sinusoidal voltage (DC offset 3.5 Vpp) to the stage at 0.4 Hz frequencies. The oscillating movement imposed on the stage highlights the indentation test and the tensile test known for the study of mechanical behavior of materials. The mechanical properties found are: the modulus of elasticity (Young Modulus), the shear modulus, the coefficient of hardening and erythrocyte resistance coefficient. The axial shear modulus 25.00 ± 1.5 μN/m and the transversal shear modulus 15.7 ± 4.63 μN/m were compared to those in the literature. These values were respectively determined by Hooke’s law and the Hertz model.
DOI
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