Higher order structure (HOS) analysis of Yersinia pestis virulence factor Caf1 polymerization using carboxyl-group foot-printing

CASSS WCBP Symposium 2026 -- Mass spectrometry carboxyl-group foot-printing has been used to investigate the higher order structure (HOS) of recombinant proteins. This involves the temporal, selective chemical modification of carboxyl groups with 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC) and glycine ethyl ester (GEE) to produce an 85 Da adduct: liquid chromatography mass spectrometry (LC-MS) peptide mapping was then used to detect and quantify the modified aspartate and glutamate residues. To test out this HOS methodology, we have applied it to look at the polymerization of the Yersinia pestis virulence factor capsular fraction 1 (Caf1), which plays a pivotal role in the pathogenicity of  Y. pestis, the causative agent of bubonic plague. Caf1 is a 15.5 KDa, thermostable protein that assembles into a highly stable polymeric capsule on the surface of Y. pestis that inhibits phagocytosis.