Discovery of novel metabolites using click-chemistry tools

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Mass spectrometry (MS)-based metabolomics is an emerging field focused on the profiling of small, naturally occurring molecules collectively known as the "metabolome". Metabolomics provides a powerful strategy to understand and investigate quantitative changes associated with a unique phenotype at the molecular level. Specifically, the relatively novel approach of untargeted metabolomics provides a comprehensive systems-wide view of metabolites or pathways with direct functional consequences. However, compared to hypothesis-driven targeted approaches, untargeted approaches are hampered by difficulties in exact compound identification, issues with limited compound abundance in samples, and low interpretability with respect to biological significance. These limitations become more challenging in the case of unknown metabolites (the "dark metabolome"), which account for ~90% of detected compounds, leaving thousands of compounds that remain to be catalogued and potentially linked to biologically significant phenotypes. My research program is aimed at discovering and mapping unknown endogenous metabolites in living organisms and determining their biosynthesis and function. Here we propose to expand the metabolome coverage and discovery in untargeted metabolomics by developing a novel systematic approach based on innovative click-chemistry metabolic labeling for metabolite enrichment. This will allow systematic mapping of target pathways, identify new and low abundance metabolites and elucidate the biosynthesis of interesting metabolites. Aim 1: Develop an optimize click-chemistry tools and resources to tag, enrich and subsequently identify novel metabolic species. Aim 2: Map the sphingolipidome in eukaryotic metabolism and annotate new metabolic species. Aim 3: Determine the substrate and products of orphan and poorly annotated enzymes. The potential impact of this research is a crucial improvement in the metabolome coverage. It will enhance our ability to explore and map metabolic pathways and discover novel metabolic species and networks. The anticipated result of this research lies in an improved methodology to specifically enrich metabolites in targeted pathways and remove interfering molecules. The impact will range from basic research and understanding of metabolism to identifying targets for therapeutical treatments.