The Discovery of Novel Metabolic Pathways for the Biosynthesis and Degradation of Complex Carbohydrates within the Human Gut Microbiome

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The primary focus of this research proposal is the discovery and elucidation of novel biochemical pathways for the biosynthesis and metabolism of complex and simple carbohydrates in the human gut microbiome. The total number of genes contained within the distinct bacterial species that inhabit the human gut exceeds the number of human genes by more than two orders of magnitude. The metabolic diversity within these bacteria contributes significantly to the maintenance of human health and physiology. Unfortunately, a significant fraction of the enzymes and metabolic pathways contained within the bacterial species localized in the human gut have an uncertain, unknown, or incorrect functional annotation. This uncertainty demonstrates that a substantial fraction of the metabolic potential found within the human gut microbiome remains to be properly characterized. The experimental approach for the discovery and elucidation of novel biochemical pathways for the metabolism of complex carbohydrates will employ the concerted and synergistic utilization of computational biology, bioinformatics, three-dimensional protein structure determination, metabolomics, and physical screening of focused compound libraries. This investigation will further be directed towards a complete understanding of the assembly and biosynthesis of the diverse capsular polysaccharides in the human pathogen Campylobacter jejuni, the leading cause of human gastroenteritis world-wide. The capsular polysaccharides are important for the invasion and colonization of the host organism and the monosaccharides that comprise the CPS in various strains of C. jejuni are unusual and complex. This endeavor will focus on the elucidation of the molecular pathways for the biosynthesis of the unusual array of monosaccharide building blocks and the associated molecular logic for the directed assembly of unique polysaccharide sequences by a series of sugar transferase enzymes. The determination of the substrate and reaction diversity contained within these newly discovered enzyme-catalyzed reactions will provide unique insights into the molecular mechanisms for the evolution and development of novel enzymatic activities and will provide potential targets for therapeutic intervention.