Structural insights into a cooperative switch between one and two FimH bacterial adhesins binding pauci- and high-mannose type N-glycan receptors

The FimH type-1 fimbrial adhesin allows pathogenic Escherichia coli to adhere to glycoproteins in the epithelial linings of human bladder and intestinal tract, by using multiple fimbriae simultaneously. Pauci-and high-mannose type N-glycans are natural FimH receptors on those glycoproteins. Oligomannose-3 and oligomannose-5 bind with the highest affinity to FimH by using the same Man alpha 1,3Man branch. Oligomannose-6 is generated from oligomannose-5 in the next step of the biogenesis of high-mannose N-glycans, by the transfer of a mannose in alpha 1,2-linkage onto this branch. Using serial crystallography and by measuring the kinetics of binding, we demonstrate that shielding the high-affinity epitope drives the binding of multiple FimH molecules. First, we profiled FimH glycan binding on a microarray containing pauci-mannosidic N-glycans and in a FimH LEctPROFILE assay. To make the transition to oligomannose-6, we measured the ki-netics of FimH binding using paucimannosidic N-glycans, glycoproteins and all four alpha-dimannosides conjugated to bovine serum albumin. Equimolar mixed interfaces of the dimannosides present in oligomannose-6 and molecular dy-namics simulations suggest a positive cooperativity in the bivalent binding of Man alpha 1,3Man alpha 1 and Man alpha 1,6Man alpha 1 dimannosides. The binding of core alpha 1,6-fucosylated oligomannose-3 in cocrystals of FimH is monovalent but interestingly the GlcNAc1-Fuc moiety retains highly flexi-bility. In cocrystals with oligomannose-6, two FimH bacterial adhesins bind the Man alpha 1,3Man alpha 1 and Man alpha 1,6Man alpha 1 end-ings of the second trimannose core (A-40-B). This cooperative switch towards bivalent binding appears sustainable beyond a molar excess of oligomannose-6. Our findings provide impor-tant novel structural insights for the design of multivalent FimH antagonists that bind with positive cooperativity.

Automated summary

The FimH type-1 fimbrial adhesin allows pathogenic Escherichia coli to adhere to glycoproteins in human bladder and intestinal tract linings. Oligomannose-3 and oligomannose-5 have the highest affinity for FimH. Our findings provide important structural insights for the design of multivalent FimH antagonists that bind with positive cooperativity.