Identification and Characterization of Novel Outer Membrane Proteins of Brachyspira pilosicoli

Abstract

Brachyspira pilosicoli is a pathogenic, Gram-negative, spirochete bacterium that causes intestinal spirochetosis (IS) in birds, pigs, and humans and is distributed worldwide. This anaerobic intestinal bacterium colonizes the large intestine, potentially leading to colitis, diarrhoea, and decreased growth rate. Outer membrane proteins of Gram-negative bacteria play crucial roles in adhesion and host-pathogen interaction, helping the bacteria to evade the host immune system, and enhancing their virulence. However, B. pilosicoli outer membrane proteins are yet to be identified and characterized. Here, we report the computational discovery of 42 outer membrane {beta}-barrel (OMBB) proteins in B. pilosicoli proteome predicted using a consensus-based computational framework. {beta}-barrel architectures of the predicted proteins were confirmed by generating AlphaFold 3-based structural models. Structure-based functional annotation predicted putative functions for the identified OMBB proteins, including BamA homolog involved in folding and membrane insertion of OMPs, LptD homolog involved in transport of lipopolysaccharides into the OM, efflux pumps, transporters, enzymes, diffusion channels, and porins. Sequence variations across nine strains of B. pilosicoli were identified and mapped onto structural models, revealing that many of the variations were present on the surface exposed loop regions of the {beta}-barrel structures. Our in-silico analysis has identified 42 OMBB proteins, including homologs of BamA, LptD, TolC, TonB-dependent receptors, CsgG. Seven of these were identified as hypothetical proteins. Computational characterization of the predicted OMBB proteins offer insights into their potential roles in physiology, virulence, and disease pathogenesis, highlighting their potential for applications in diagnostics, vaccine development, or therapeutic interventions.