The main goal of this competing renewal proposal is to study how shear force that is created by the fluid flow affects the uropathogenicity-relevant functional properties of the Escherichia coli fimbriae, withprimary focus on the mannose-sensitive, type 1fimbrial adhesin, FimH. Analysis of naturalvariants of FlmH protein has been the focus of our current grant that led us to the finding that strength of the FimH-mediated mannose- binding is dramatically enhanced by the presence of shear stress. We demonstrated that tensile mechanical force is likely to induce specific conformational changes in the FimH protein that are associated with the shear-dependent binding phenotype. In this renewal proposal we intend to use different shear-associated variants of FimH to understand possible roles of the shear-enhanced mode of the FimH-mediated adhesion that is relevantto colonization of the urinarytract by E, coli. We will perform functional analysis of various naturally-occurring FimH variants and determine binding shear-spectrum of wild-typeE. coli strains and FimH-functionalized beads. To characterize physiological significanceof the FimH shear-associated phenotypes we also will determine FimH-mediated binding to natural-likereceptors and variable shear conditions; adhesion and invasion of uroepithelial cells, and colonization in animal model of urinary tract infection. In addition, we will characterize inhibitorsof the FimH-mediated bindingunder different shear. Studies proposed here will serve as model ones to provide a spectrum of novel information regarding shear effects on bacterial adhesion, particularlyin connection to the bacterial pathogenesis.
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