Developing glycan-directed tools to investigate microbial infection

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7. Project Summary/Abstract Glycans and glycoconjugates coating the surface of bacteria underscore critical roles in various biological processes. One such process, which currently lacks mechanistic knowledge, is the interaction of the N-linked heptasaccharide from enteric pathogen Campylobacter jejuni with host cell receptors, resulting in human infection. N- and O-glycosylation systems in other pathogenic bacteria have shown similar essential interactions to maintain virulence. Therefore, understanding these glycan-receptor interactions will provide information on the molecular mechanisms behind microbial infection and reveal insights for future therapeutic intervention. Current biochemical tools lack appropriate characteristics to study host-pathogen interactions, and there is a resulting demand for the development of glycan-directed tools which can be leveraged to elucidate critical molecular determinants of pathogenesis. The proposed project focuses on the generation of unique biochemical species to understand the role of N-glycosylation in C. jejuni infection. Specifically, these include glycan- decorated magnetic beads, multivalent glycan-functionalized peptides and glycan-specific protein binders. The two multivalent glycan scaffolds will be attained through bioconjugate chemistries and chemoenzymatic synthesis and the resulting chemically-defined scaffolds can afford specific protein binders through a yeast surface display directed evolution. All three species will be applied in a microphysiological gastrointestinal model (GutChip) of C. jejuni infection to afford pathogenic mechanistic information. Glycan-coated magnetic beads interact with host cell receptors, allowing for the elucidation of the specific protein which binds glycans. Multivalent glycan-functionalized polymers of heptasaccharide fragments can compete for host cell receptors, providing information on the C. jejuni glycan regions involved in pathogenesis. The evolved protein binders may mask the exposed N-glycan, thwarting glycan-mediated infectious mechanisms. The lessons learned from the C. jejuni model will be broadly-applicable to other bacterial threats and therefore invaluable to the glyco- and microbiology communities. This project will be performed under the sponsorship of Professor Barbara Imperiali, in the Departments of Biology and Chemistry at Massachusetts Institute of Technologies. Her support, the lab’s expertise, and the wealth of resources available at MIT will allow for the successful completion of the proposed research project.