Autotransporter proteins (AT) represent the biggest family of secreted and outer membrane proteins in Gram-negative bacteria. They promote phenotypes such as cell adhesion and toxicity as well as cell aggregation and biofilm formation, which allow bacteria to resist the effect of antibiotics and immune responses from hosts. The predominant group in the AT family are the surface proteins known as the AIDA-I type adhesins. Our research focuses on the most prevalent adhesin, Antigen 43 (Ag43) from uropathogenic Escherichia coli. We aim at investigating the mechanism of action and processing of this AT and analysing the structure-function relationships of Ag43 homologues.
Combining X-ray crystallography, biophysics and cell based assays, we have shown that the functional α-domain of Ag43b from E. coli CFT073 folds into a three-stranded beta-helix, mimicking the previously characterised Ag43a. We have also determined the mechanism of self-association of this AT, which differs slightly from that of Ag43a, explaining their different function.
Additionally, this project aims to identify molecules that block the function of Ag43. This has been attempted using single-domain antibodies derived from the variable region (vNAR) of a shark antibody known as the IgNAR. Screening of a vNAR phage-display library resulted in the identification of two different binders that recognise three Ag43 homologues. Ongoing work with these vNARs will establish their ability to halt the aggregative ability of Ag43.
The outcomes of this work will provide essential data for future studies on the function and inhibition of this key AT.