Deciphering the Mechanical Properties of Type III Secretion System EspA Protein by Single Molecule Force Spectroscopy

被引:7
作者
Nadler, Hila [1 ]
Shaulov, Lihi [2 ]
Blitsman, Yossi [1 ]
Mordechai, Moran [1 ]
Jopp, Jurgen [3 ]
Sal-Man, Neta [2 ]
Berkovich, Ronen [1 ,3 ]
机构
[1] Ben Gurion Univ Negev, Dept Chem Engn, IL-8410501 Beer Sheva, Israel
[2] Ben Gurion Univ Negev, Dept Microbiol Immunol & Genet, IL-8410501 Beer Sheva, Israel
[3] Ben Gurion Univ Negev, Ilse Katz Inst Nanoscale Sci & Technol, IL-8410501 Beer Sheva, Israel
基金
以色列科学基金会;
关键词
ENTEROPATHOGENIC ESCHERICHIA-COLI; COILED-COIL PEPTIDES; TITIN; RESISTANCE; DOMAIN; ELASTICITY; FILAMENTS; DYNAMICS; DNA; INTERMEDIATE;
D O I
10.1021/acs.langmuir.8b01198
中图分类号
O6 [化学];
学科分类号
0703 ;
摘要
Bacterial pathogens inject virulence factors into host cells during bacterial infections using type III secretion systems. In enteropathogenic Escherichia coli, this system contains an external filament, formed by a self-oligomerizing protein called E. coli secreted protein A (EspA). The EspA filament penetrates the thick viscous mucus layer to facilitate the attachment of the bacteria to the gut epithelium. To do that, the EspA filament requires noteworthy mechanical endurance considering the mechanical shear stresses found within the intestinal tract. To date, the mechanical properties of the EspA filament and the structural and biophysical knowledge of monomeric EspA are very limited, mostly due to the strong tendency of the protein to self-oligomerize. To overcome this limitation, we employed a single molecule force spectroscopy (SMFS) technique and studied the mechanical properties of EspA. Force extension dynamic of (I91)(4)-EspA-(I91)(4) chimera revealed two structural unfolding events occurring at low forces during EspA unfolding, thus indicating no unique mechanical stability of the monomeric protein. SMFS examination of purified monomeric EspA protein, treated by a gradually refolding protocol, exhibited similar mechanical properties as the EspA protein within the (I91)(4)-EspA-(I91)(4) chimera. Overall, our results suggest that the mechanical integrity of the EspA filament likely originates from the interactions between EspA monomers and not from the strength of an individual monomer.
引用
收藏
页码:6261 / 6270
页数:10
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