Recruitment and Mechanism of the Bacterial Type IV Pilus Motors

The type IVa pilus system can be used for adhesion, natural competence, phage adsorption, folded-protein secretion, surface sensing, and surface associated motility. Likewise, it is a major virulence factor in many bacterial pathogens. This system extends and retracts a thin and long protein fibre from the bacterial inner membrane. PilB and PilT are the cytoplasmic ATPases that power pilus extension and retraction, respectively. PilC is an inner membrane protein that bridges the cytoplasmic ATPases to the pilus. PilM is a cytoplasmic protein that is predicted to recruit PilB or PilT, and also binds to the cytoplasmic N-terminal residues of PilN, another inner membrane protein. Herein crystal structures of PilM alone and bound to the N-terminal residues of PilN demonstrate that PilN-binding causes structural changes in PilM. PilM bound to PilB and PilT, and evidence was provided that PilN-binding to PilM inversely modulates these interactions. This is consistent with a model in which PilM recruits PilB or PilT to decide whether the pilus system extends or retracts. The crystal structures of the core ATPase domains of PilB bound to ADP and a non-hydrolyzable ATP analogue were determined. Analysis of the differences between structures permitted the deduction of the catalytic mechanism and domain movements of PilB, which with PilC would support the assembly of a right-handed helical pilus. Extrapolating this analysis to a previously published structure of PilT suggests a mechanism for how PilT would enable right-handed pilus disassembly. However, the available crystal structures of PilT, as well as new structures determined herein, are heterogeneous and not obviously consistent with the structure used for this analysis. CryoEM analysis revealed PilT, but not PilB, has preferences for conformation that are obscured by crystal lattice formation. The physiologic importance of these conformations was validated, allowing for the proposal of the most comprehensive model of PilT function to date.

Department dc:contributor.department

Biochemistry

Year dc:date.issued

2019

Author dc:creator

• McCallum, Matthew

Advisor dc:contributor.advisor

• Howell, P. Lynne

• PilB
• PilM
• PilN
• PilT
• pilus
• T4P

Handle dc:identifier.uri

http://hdl.handle.net/1807/108216

OAI identifier oai:identifier

oai:utoronto.scholaris.ca:1807/108216