Genome sequencing reveals complex secondary metabolome in the marine actinomycete Salinispora tropica

Abstract

Recent fermentation studies have identified actinomycetes of the marine-dwelling genus Salinispora as prolific natural product producers. To further evaluate their biosynthetic potential, we sequenced the 5,183,331-bp S. tropica CNB-440 circular genome and analyzed all identifiable secondary natural product gene clusters. Our analysis shows that S. tropica dedicates a large percentage of its genome ( approximately 9.9%) to natural product assembly, which is greater than previous Streptomyces genome sequences as well as other natural product-producing actinomycetes. The S. tropica genome features polyketide synthase systems of every known formally classified family, nonribosomal peptide synthetases, and several hybrid clusters. Although a few clusters appear to encode molecules previously identified in Streptomyces species, the majority of the 17 biosynthetic loci are novel. Specific chemical information about putative and observed natural product molecules is presented and discussed. In addition, our bioinformatic analysis not only was critical for the structure elucidation of the polyene macrolactam salinilactam A, but its structural analysis aided the genome assembly of the highly repetitive slm loci. This study firmly establishes the genus Salinispora as a rich source of drug-like molecules and importantly reveals the powerful interplay between genomic analysis and traditional natural product isolation studies.

Fig. 1.

Circular chromosome of S. tropica CNB-440, oriented to the dnaA gene. The outside outer ring shows the locations of secondary metabolic gene clusters. The inside outer ring shows the locations of putative mobile genetic elements. The center ring shows a normalized plot of GC content (maximum, 75.5%; minimum, 60.3%; average, 69.5%). The inner ring shows a normalized plot of GC skew (maximum, 0.2346; minimum, −0.2504; average, −0.0020).

Fig. 2.

Selected genes from S. tropica modular biosynthetic enzyme systems and their associated natural products. (A–D) Putative siderophore clusters. (A) sid1 cluster in comparison to the desferrioxamine cluster from Streptomyces coelicolor A3 (2). (B) sid2 cluster in comparison to the yersiniabactin cluster from Y. pestis. (C) sid3 cluster with putative product intermediate. (D) sid4 cluster in comparison to proposed S. coelicolor‘coelibactin' cluster (unconfirmed structure). (E–G) Novel modular enzyme systems. (E) salAB from proposed salinosporamide cluster. (F) lymAB from proposed lymphostin cluster. (G) slmNABCD modular PKS system from proposed salinilactam cluster (stereo chemistry proposed, green portion is derived from the proposed lysine-based starter unit). Domain notation: A, adenylation (amino acid substrate noted); C, condensation; Ccyc, condensation with cyclization; PCP, peptidyl carrier protein; KS, β-ketoacyl synthase; AT, acyl-CoA/ACP transacylase (activates malonyl-CoA unless otherwise noted); mAT, methylmalonyl-CoA/ACP transacylase; DH, dehydratase; ACP, acyl carrier protein; TE, thioesterase; OR, oxidoreductase; MT, SAM-dependent methyltransferase; NAT, N-acetyl transferase; unk, unknown.

Fig. 3.

Enediyne polyketide biochemistry in S. tropica. (A) Representative enediyne structures. (B) Phylogenetic tree of representative enediyne PKSs. GenBank accession nos.: MadE, AAQ17110; NcsE, AAM78012; SgcE, AAL06699; CalE8, AAM94794; EspE, AAP92148; DynE, AAN79725. (C) Proposed biosynthesis of sporolide A.