Nocardiopyrone A is a structurally novel natural product originally isolated from the marine-derived Nocardiopsis sp. CMB-M0232 actinomycete bacterium. The cluster of genes necessary for the biosynthesis of this natural product is unknown. The sequenced genome of this marine bacterium revealed a ~25-kb gene cluster encoding enzymes with hypothesized roles in nocardiopyrone assembly. To probe the role of this gene cluster in nocardiopyrone biosynthesis, the cluster was cloned into a vector and introduced into Streptomyces lividans TK24 as a heterologous expression host. Additionally, knockout mutants of predicted polyketide synthase, crotonyl CoA reductase, and 3-hydroxybutyryl CoA dehydrogenase-encoding genes from that cluster were constructed using PCR-targeted gene replacement. S. lividans cultures featuring the complete gene cluster and knockout mutants were grown for seven days alongside a control culture lacking all of these biosynthetic genes. These cultures were chemically extracted and metabolic profiles were obtained using high performance liquid chromatography (HPLC), and compared between all cultures. The elimination of the crotonyl CoA reductase-encoding gene resulted in accumulation of a metabolite at a retention time slightly less than that expected of nocardiopyrone. Isolation of this metabolite will be required to determine if it is a precursor of nocardiopyrone A that accumulates in response to deletion of the crotonyl CoA reductase gene.

Evaluation of the putative nocardiopyrone biosynthetic gene cluster from a marine-derived actinomycete

251st National Meeting (2016)

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