Predicting Homogeneous Pilus Structure from Monomeric Data and Sparse Constraints.

Area of Science:

• Microbiology
• Structural Biology
• Computational Biology

Background:

• Type IV pili (T4P) and Type II Secretion System (T2SS) pseudopili are crucial extracellular protein filaments in bacteria.
• These pili are assembled from homologous subunits known as pilins.
• Understanding the atomic structure of these filaments is essential for elucidating their diverse biological roles.

Purpose of the Study:

• To develop and validate a new computational approach for predicting pseudo-atomic models of microbial pili and pseudopili.
• To assess the flexibility and conformational variability of pilus structures.

Main Methods:

• A novel modeling approach combining ambiguous symmetric constraints with sparse experimental distance information.
• Validation using experimental data from Neisseria gonorrhoeae (GC pilus), Vibrio cholerae (TCP), and Klebsiella oxytoca (PulG pilus).
• Computational error analysis and global sampling to explore conformational space.

Main Results:

• Successfully reconstructed pseudo-atomic models for GC pilus, T2SS pseudopilus, and T4b pilus.
• Demonstrated that pilus subunits exhibit slight flexibility, challenging the rigid-body assumption.
• Identified that pili can adopt multiple, but not an unlimited number of, intact conformations.

Conclusions:

• The developed method provides a powerful tool for structural prediction of pili and pseudopili without relying on EM maps or precise symmetry.
• Pilus structural models should account for subunit flexibility and potential conformational heterogeneity.
• Further structural studies are warranted to fully explore the conformational landscape of bacterial filaments.