ROLE OF WAAL LIGASES IN SERUM RESISTANCE OF YERSINIA ENTEROCOLITICA SEROTYPES O:3 AND O:8

Abstract

The aim of current study was to estimate WaaL ligase contribution in lipopolysaccharide (LPS) phenotype profile formation of Y. enterocolitica serotype O:3 (YeO3) and O:8 (YeO8) bacteria and its participation in serum killing protection. In lipopolysaccharide (LPS) biosynthesis of Gram-negative bacteria the waaL-encoded ligase joins O-polysaccharide (O-Ag) and outer core (OC) onto lipid A-core oligosaccharide. Three waaL genes named as waaLOS, waaLPS and waaLXS were identified from Yersinia enterocolitica genome. Methods. The waaL-knock-out mutants were created by allelic exchange strategy. The LPS phenotypes of created mutants were visualized by silverstained DOC-PAGE and immunoblotting with specific outer core (core oligosaccharide, hexasaccharide, OC) and O-polysaccharide (OPS or O-Ag) monoclonal antibodies. To study the contribution of WaaLOS and WaaLPS to the survival of Yersinia bacteria in non-immune human serum, we constructed the series of single and double ligase mutants. Survival of bacteria was analyzed in normal serum (with functional classical, lectin, and alternative complement activation pathways) and EGTA-Mg-treated serum (only alternative pathway functional). Results. Our results demonstrated that WaaL ligases participate in the synthesis of proper LPS structure and play an important role in protection against serum killing. Conclusions. The LPS ligases of YeO3 exhibit relaxed donor substrate specificity. Under given conditions the effect of WaaLOS ligase is more significant for OC and OPS ligation onto lipid A then WaaLPS one.