Numerous bioactive secondary metabolites contain amino acids that are not found in proteins. These unique building blocks may be biosynthesized by discrete pathways or they may be elaborated from the 20 canonical amino acids.  The ability of microbes to generate novel amino acids can greatly expand the chemical space and reactivity of natural products and can have a profound effect on biological activity. I will present two studies that are focused on amino acids that are incorporated into bioactive secondary metabolites; the first example involves the biosynthesis of an unnatural amino acid, and the second example is focused on the derivatization of tryptophan.The nonproteogenic amino acid 3,5-Dihydroxyphenylglycine (DPG) is synthesized and incorporated into the vancomycin family of antibiotics.  A key enzyme in this pathway is DpgC, which performs a 4-electron oxidation without the use of metals or cofactors. We studied this unique enzyme using the tools of synthetic organic chemistry, enzymology and structural biology. The structure of DpgC was determined with a bound synthetic inhibitor, which allowed us to identify the active site. We determined the mechanism of DpgC via a systematic study of mutations of catalytically relevant amino acids and alternate substrates.  Lymphostin is a member of the pyrroloquinoline alkaloid family of natural products that contain a tryptophan-derived tricyclic core. Herein we describe the biosynthetic basis of lymphostin production in the marine microbe Salinispora. This pathway involves a uniquely organized polyketide synthase-nonribosomal peptide synthetase hybrid, which incorporates a modified tryptophan that originates from a precursor peptide. We are using a variety of genetic and biochemical techniques to elucidate the method in which this microbe extracts, modifies, and incorporates this amino acid into the final natural product.  The knowledge gained from this study can lead to the discovery of pathways for the larger class of pyrroloquinoline alkaloids, which have primarily been extracted from marine sponges or mushrooms.

Natural and nonproteogenic amino acids in natural product biosynthesis

244th National Meeting (2012)

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