Investigating biosynthetic enzymes to enhance natural product discovery

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Project Summary Natural products (NPs) have diverse complex structures and play a vital role in drug discovery and development. Although the rate of discovery of natural products has increased significantly, traditional bioactivity-guided discovery methods frequently lead to the re-discovery of known compounds. Therefore, genetic information- driven isolation is becoming a powerful tool to uncover novel NPs. Genome mining efforts that target genes responsible for the biosynthesis of core structures of major NP classes, has identified tens of thousands of new biosynthetic gene cluster (BGC) families predicted to produce novel compounds. While this method has been very successful in predicting compounds from these major classes, it is limited to those scaffolds and cannot identify other specific features of NP structures. Furthermore, these methods will overlook “hidden” BGCs that do not contain traditional core biosynthetic machinery, leaving a major gap in NP discovery. The first direction of this proposal aims to use specialized genome mining strategies that target NP glycosyltransferases to identify uncharacterized BGCs that produce bioactive glycosylated NPs with distinct core structures. Consequently, identified compounds will be directly linked to their BGC and likely have biosynthetic pathways that consist of unique enzymes and biochemical reactions. The second direction of this proposal is to interrogate the activities and mechanisms of new biosynthetic enzymes. These will include enzymes discovered in direction one, along with enzymes responsible for the biosynthesis of the aminocyclitol found in hygromycin A, that contains a rare modification essential for bioavailability. Subsequent re-integration of newly characterized biosynthetic enzymes into standard or specialized genome mining methods will assist in annotating and identifying additional BGCs to improve NP discovery. In addition, these enzymes will add to the growing toolbox of biocatalytic reactions exploited for unnatural small molecule biosynthesis. Ultimately, this research program will significantly advance NP and enzymology research to boost drug discovery and development for the benefit of human health.