Identification of a biosynthetic gene cluster for the polyene macrolactam sceliphrolactam in a Streptomyces strain isolated from mangrove sediment

Abstract

Streptomyces are a genus of Actinobacteria capable of producing structurally diverse natural products. Here we report the isolation and characterization of a biosynthetically talented Streptomyces (Streptomyces sp. SD85) from tropical mangrove sediments. Whole-genome sequencing revealed that Streptomyces sp. SD85 harbors at least 52 biosynthetic gene clusters (BGCs), which constitute 21.2% of the 8.6-Mb genome. When cultivated under lab conditions, Streptomyces sp. SD85 produces sceliphrolactam, a 26-membered polyene macrolactam with unknown biosynthetic origin. Genome mining yielded a putative sceliphrolactam BGC (sce) that encodes a type I modular polyketide synthase (PKS) system, several β-amino acid starter biosynthetic enzymes, transporters, and transcriptional regulators. Using the CRISPR/Cas9–based gene knockout method, we demonstrated that the sce BGC is essential for sceliphrolactam biosynthesis. Unexpectedly, the PKS system encoded by sce is short of one module required for assembling the 26-membered macrolactam skeleton according to the collinearity rule. With experimental data disfavoring the involvement of a trans-PKS module, the biosynthesis of sceliphrolactam seems to be best rationalized by invoking a mechanism whereby the PKS system employs an iterative module to catalyze two successive chain extensions with different outcomes. The potential violation of the collinearity rule makes the mechanism distinct from those of other polyene macrolactams.

Dark, rare and inspirational microbial matter in the extremobiosphere: 16 000 m of bioprospecting campaigns

“…While recognizing that more mundane environments can contain novelty for the bioprospector (e.g. forest soil [73], street sediments [74]) we opine that the case for prioritizing extreme biomes is sound as it moves bioprospecting action away from well-probed terrestrial streptomycetes towards microbial communities that have evolved under harsh selective conditions during the course of which novel biochemistry has been generated [75]. As Rateb et al report [66], a total of 46 natural products representing diverse chemical classes have been isolated from Atacama Desert actinobacteria during the past 7 years.…”

Genome Mining Revealed a High Biosynthetic Potential for Antifungal Streptomyces sp. S-2 Isolated from Black Soot

“…Both groups of BGCs are known to encode proteins involved in biosynthesis and the processing of secondary metabolites with various antibiotic activities [65]. In other microorganisms, NRPS are responsible for synthesis of compounds such as actinomycin, vancomycin, and coelibactin [66][67][68], and T1PKS produce compounds such as rapamycin, cyphomycin, and glutaramides [18,25,69]. On the other hand, phenotype fingerprinting using a microplate assay showed that the S-2 strain was susceptible to several antibacterial antibiotics targeting protein synthesis.…”

Characterization of Cas proteins for CRISPR‐Cas editing in streptomycetes

“…Testing multiple protospacers may also help when no genome editing is observed. Low and coworkers showed that only one out of five protospacers yielded the desired deletion of a polyketide synthase module in Streptomyces Sd‐85 (Low et al, ). Optimizing Cas expression through various strategies to balance transcription and translation efficiencies has been successful in alleviating nuclease‐induced toxicity in organisms, such as mycobacteria, cyanobacteria, and corynebacteria (Cho et al, ; Sun et al, ; Wendt, Ungerer, Cobb, Zhao, & Pakrasi, ).…”

“…Notably, sequence‐specific toxicity is observed for the catalytically dead spCas9 (dCas9) in Escherichia coli and can be alleviated by reducing dCas9 expression levels (Cui et al, ). In other contexts, even after a successful introduction of recombinant CRISPR‐Cas systems, no editing is observed in exconjugants expressing spCas9 and its guide RNA (Low et al, ). In E. coli , modifications induced by CRISPR‐Cas9 double‐stranded breaks are circumvented with RecA‐dependent repair (Moreb et al, ).…”