MIRA: Probing Glycan Polymer Patterns on Bacterial Cell Surfaces

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Project Summary/Abstract Our planet is inhabited by trillions of bacteria that live inside and outside of humans. The “skin”, or surface, of bacteria is called the cell envelope, and functions to separate us from them. Although some bacteria are symbionts, infection by pathogenic bacteria is still a major cause of death worldwide. While Gram-negative bacteria contain a protective outer membrane layer absent in most Gram-positives, almost all bacteria contain polymers composed of unique patterns of glycans that extend from the cell surface. Bacterial surface sugar polymers, or exo-polysaccharides, act as molecular barcodes that distinguish different strains of bacteria within a single species. Many bacterial exo-polysaccharides contain rare sugars, which are monosaccharides that are absent in other organisms, including humans. While exo-polysaccharides are necessary for host infection, we still lack an understanding of how rare sugar-containing glycan polymers are assembled, recognized, and enable survival in the host. My laboratory seeks to generate chemical and biochemical tools to study bacterial protein and glycan pathways that enable survival in different environments. Our main areas of focus are: (1) development of small molecule regulators of bacterial chaperone function; (2) manipulation of cell surface sugar patterns to selectively label and disable bacteria. This proposal focuses on the latter program, in which we identify rare saccharide subunits that are unique to Gram-negative cell surface polymers called O-antigens, and represent key epitopes that mediate interactions with hosts and susceptibility to antibiotics. Over the next five years, we will address the following questions: (1) Can we improve chemoenzymatic routes to rare sugar precursor substrates? (2) How do glycosyltransferases recognize rare sugar substrates to build O-antigens? (3) Are O-antigen glycosyltransferases regulated via protein-protein interactions? (4) What host protein structural motifs are involved in bacterial rare sugar recognition? (5) Can we identify new host proteins involved in bacterial recognition? To answer these questions, we will use a multidisciplinary approach, involving a combination of organic chemistry, chemical biology, biochemistry, microbiology and sequencing-based analyses. This work will significantly expand our understanding of cellular mechanisms underlying bacterial polysaccharide synthesis, and will teach us how humans recognize foreign sugars. Relevance to public health: In addition to providing fundamental insight into the production of bacterial factors that are important for infection, the results of this proposal will inform novel strategies to disable hard-to-treat Gram-negative infections by interference of essential host-pathogen interactions, as well as biomolecular reagents to recognize bacterial oligosaccharide structures for new diagnostics.