Identification of Protein Domains on Major Pilin MrkA that Affects Mechanical Properties of Klebsiella pneumoniae Type 3 Fimbriae

Chia-Han Chan, FENG-JUNG CHEN, Ying-Jung Huang, Shin-Yu Chen, Kuo-Liang Liu, Zhe-Chong Wang, Hwei-Ling Peng, Tri-Rung Yew, Cheng-Hsien Liu, Gan-Guang Liou, Ken Y. Hsu, Hwan-You Chang, and Long Hsu

The Klebsiella pneumoniae type 3 fimbriae are mainly composed of MrkA pilins that assemble into a helix-like filament. This study determined the biomechanical properties of the fimbriae and analyzed 11 site-directed MrkA mutants to identify domains critical for the properties. Escherichia coli strains expressing type 3 fimbriae with an Ala substitution either at F34, V45, C87, G189, T196, or Y197 resulted in a significant reduction in biofilm formation. The E. coli strain expressing MrkAG189A remained capable of producing a normal amount of fimbriae. Although F34A, V45A, T196A, and Y197A substitutions expressed on E. coli strains produced sparse quantities of fimbriae, no fimbriae were observed on the cells expressing MrkAC87A. Further investigations of the mechanical properties of the MrkAG189A fimbriae with optical tweezers revealed that, unlike the wild-type fimbriae, the uncoiling force for MrkAG189A fimbriae was not constant. The MrkAG189A fimbriae also exhibited a lower enthalpy in the differential scanning calorimetry analysis. Together, these findings indicate that the mutant fimbriae are less stable than the wild type. In conclusion, this study has demonstrated that the C-terminal β-strands of MrkA are required for the assembly and structural stability of fimbriae.

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