Deciphering Helicobacter pylori's glycocode: uncovering and harnessing drug targets - Antibiotic resistant bacterial pathogens cause more than 2.8 million new infections and 32,000 deaths each year in the US,1 indicating the ineffectiveness of our existing arsenal of antibiotics. Even when antibiotics are effective at eradicating infection, most are inherently nonspecific for pathogens and have unintended conse- quences on beneficial microbiota.2 Bacterial cell surface glycans are quintessential drug targets due to their critical role in colonization of the host, pathogen survival, and immune evasion.3-5 Despite the importance of glycan biosynthesis as a virulence factor, the systematic study and inhibition of bacterial glycans remains chal- lenging due to their utilization of rare deoxy amino sugars and their production of products that are refractory to traditional glycan analysis. The deployment of chemical tools to study bacterial glycans is a crucial step toward developing new glycosylation-based strategies to eradicate pathogenic infections. My laboratory seeks to apply metabolic oligosaccharide engineering (MOE) tools and develop new tools to study and perturb glycan biosyn- thesis in the gastric pathogen Helicobacter pylori (Hp). The central hypothesis of the application is that MOE with unnatural monosaccharides, including rare bacterial monosaccharide reporters,11 metabolic glycan inhibi- tors, and novel photocrosslinking sugars, will reveal the role of Hp glycans in mediating adhesion to host cells, the identity of proteins on host cells that engage in binding interactions with Hp glycans, and the role of Hp gly- cans in oncogenesis. Our hypothesis has been formulated on the basis of strong preliminary data produced in my laboratory, including the use of MOE to install detectable reporters into bacterial glycans,9-11 thus permitting the discovery of a protein glycosylation system,12, 13 the identification of glycosylation genes,14 the development of metabolic glycan inhibitors,15 and the modulation of the host immune response. The proposed work will re- veal molecular-level details about glycan-mediated host-pathogen interactions, resulting in innovative ap- proaches to treat bacterial infection. The central hypothesis will be tested by pursuing three specific aims. In Aim 1, an ordered collection of glycosylation mutants and metabolic inhibitors that disrupt glycan biosynthesis will be used to probe the role of Hp glycans in adhesion to host cells, revealing the precise roles of different classes of Hp glycans in host-pathogen interactions. In Aim 2, novel photocrosslinking bacterial sugars will be used to label Hp glycans, allowing covalent trapping, enrichment, and identification of proteins on host cells that interact with Hp glycans. In Aim 3, metabolic glycan labeling and glycoproteomics will be used to pinpoint glycan changes that occur in oncogenic Hp, and genetic and small molecule approaches will be used to re- store Hp glycans to a symbiotic state. The proposed work will yield insight into which Hp glycans mediate host cell binding and recognition, the identities of host cell receptors that bind to Hp glycans, and the structural changes that Hp glycans undergo upon bacterial transformation to an oncogenic state. Broadly, this work will introduce approaches to study and harness glycans of priority pathogens to create urgently needed antibiotics.
