The autotransporter family is the largest group of secreted and outer-membrane proteins in Gram-negative bacteria. These proteins perform a vast array of functions linked to pathogenesis, from adhesion and invasion of human host cells to the formation of cell aggregates and biofilms on biotic and abiotic surfaces. The self-associating autotransporters (SAATs) are a sub-group of autotranporters widespread across pathogens . These proteins promote the formation of aggregated communities and biofilms, which facilitate host colonization and bacterial persistence in different environmental niches.
We previously elucidated the mechanism by which the SAAT Antigen43 from uropathogenic E. coli (UPEC) promotes bacterial aggregation/biofilm formation, by means of self-association between neighbouring cells . We sought to determine if all SAATs shared a common mechanism for facilitating bacterial aggregation/biofilm formation, if this function was regulated and if it could be inhibited. TibA is a multifunctional SAAT from enterotoxigenic E. coli (ETEC), the leading bacterial cause of diarrhea. This surface protein was known to be glycosylated by the cognate glycosyltransferase TibC. We determined the crystal structures of the glycosylated and unglycosylated forms of TibA and used this to inform further biophysical and phenotypic studies. We found that TibA self-associates in a head-to-tail manner with an extensive interface, to facilitate bacterial aggregation/biofilm formation. Glycosylation by TibC was found to physically block TibA self-association to reduce bacterial aggregation/biofilm formation. Our comprehensive structural and functional analysis provide a molecular understanding of how a post-translational modification switches the activity of TibA from an aggregative molecule to and adhesin and invasin. This may represent a general mechanism for bacteria to regulate the virulence functions of the vast number of SAAT expressed on their cell surface.
We have also developed a nM inhibitor of SAAT mediated aggregation/biofilm formation and have determined the first autotransporter-inhibitor crystal structure. Molecules that block bacterial cell clusters and biofilms could be used in synergy with antibiotics, detergents or anti-biofilm agents to improve their efficacy, which would impact environmental, industrial, and human medical microbiology.