Identifying Salinispora Orphan Biosynthetic Pathways by Direct cloning and Heterologous Expression

The marine actinomycete genus Salinispora has proven to be a remarkably prolific source of structurally diverse and biologically active secondary metabolites. These compounds span virtually all known biosynthetic classes, including the proteasome inhibitor salinosporamide A that is currently in clinical trials for the treatment of cancer. Recent genome sequencing efforts have revealed that only a small fraction of the natural product biosynthetic potential from Salinispora species has been discovered. Here we report a platform for the capture of Salinispora gene clusters using a Transformation-Associated Recombination (TAR) strategy coupled with direct expression in a newly developed marine actinobacterial host. A ~25 kb noncanonical polyketide synthase (PKS) and non-ribosomal peptide synthetase (NRPS) hybrid gene cluster from Salinispora pacifica CNS863, for which the products could not be predicted, was selected, captured and heterologously expressed in the host Salinispora tropica CNB4401. The metabolites produced were isolated and structurally elucidated and represent a group of previously known bacterial fatty acid inhibitors sharing an unusual thiotetronic acid of unknown gene origin. This platform represents a streamlined method for the discovery and biosynthetic characterization of natural products from marine Salinispora species.