Structural and functional characterization of colonization factors AIBI-CS6 and AIIBII-CS6 of enterotoxigenic...

Over time, the structure and function of the broadly dispersed colonization factor (CF) CS6 of enterotoxigenic Escherichia coli (ETEC) have become more significant. CS6 is composed of tightly-associated subunits, CssA and CssB which due to presence of natural point mutation gave rise to CS6 subtypes. In contrast to the other ETEC CFs, CS6 is an afimbrial, spherical-shaped oligomers of (CssA-CssB)_n complex where 'n' is concentration dependent. In this study, we have compared AIBI-CS6 and AIIBII-CS6 structurally and functionally. The M_w of CssAI was 18.5 kDa but M_w of CssAII was 15.1 kDa. Both CssBI and CssBII had M_w of 15.9 kDa. The substitution of Gly³⁹ with Ala³⁹ in CssAI leads to reduction in Mw from 18.5 to 15.1 kDa. Due to higher M_w of CssAI, the size of AIBI concentration-dependent oligomers should be higher. However, the M_w of AIIBII oligomers were higher and AIIBII also showed higher oligomeric forms compared to AIBI both in native PAGE and electron microscopy. The oligomers of both subtypes could withstand greater temperatures and denaturant concentrations. In terms of cellular response, the levels of inflammatory cytokines were significantly higher in case of AIBI-CS6 expressing ETEC as compared to AIIBII-CS6 expressing ETEC both in vitro and in vivo. When inflammatory cytokines were evaluated after infecting suckling mice with these ETEC strains, the results were consistent. In conclusion, even though there was subtle structural difference between AIBI-CS6 and AIIBII-CS6 due to natural point mutations but ETEC strains expressing these subtypes displayed great variability in pathogenicity.