Cloning and characterization of the goadsporin biosynthetic gene cluster from Streptomyces sp. TP-A0584

Abstract

The biosynthetic gene cluster of goadsporin, a polypeptide antibiotic containing thiazole and oxazole rings, was cloned from Streptomyces sp. TP-A0584. The cluster contains a structural gene, godA, and nine god (goadsporin) genes involved in post-translational modification, immunity and transcriptional regulation. Although the gene organization is similar to typical bacteriocin biosynthetic gene clusters, each goadsporin biosynthetic gene shows low homology to these genes. Goadsporin biosynthesis is initiated by the translation of godA, and the subsequent cyclization, dehydration and acetylation are probably catalysed by godD, godE, godF, godG and godH gene products. godI shows high similarity to the 54 kDa subunit of the signal recognition particle and plays an important role in goadsporin immunity. Furthermore, four goadsporin analogues were produced by site-directed mutagenesis of godA, suggesting that this biosynthesis machinery is used for the heterocyclization of peptides.

In vitro activity of the nisin dehydratase NisB

“…The LanB dehydratases are 115-120 kDa proteins that do not share homology with the dehydratases from classes II-IV lanthipeptides or other proteins in the databases. Genes encoding homologous LanB-like enzymes are also present in the gene clusters of nonlantibiotic peptide natural products, such as the thiopeptides (16)(17)(18)(19) and goadsporin (20). These proteins are also believed to be dehydratases.…”

Biosynthesis of Heterocyclic Antibiotics in Actinomycetes and an Approach to Synthesize the Natural Compounds

“…(iv) Goadsporin immunity in the producing strain 24) The godI gene is responsible for goadsporin immunity. S. lividans harboring godI is resistant to goadsporin, and the godI-disrupted mutant of TP-A0584 is sensitive to goadsporin (Fig.…”

“…(v) An approach to synthesize the natural goadsporin analogs 24) The versatility of the goadsporin biosynthesis machinery has been indicated by the production of goadsporin analogs. The amino acid sequence of godA was changed by sitedirected mutagenesis, and four analogs were isolated from the recombinant gene strains.…”

“…(ii) Cloning of goadsporin biosynthetic gene cluster 24) It was presumed that goadsporin is ribosomally biosynthesized, and the precursor sequence is ATVSTILCSG-GTLSSAGCV; therefore, the goadsporin biosynthetic gene cluster was cloned from the TP-A0584 genomic library by using a synthetic oligonucleotide probe, which is designed from the amino acid sequence of the goadsporin precursor. Two positive clones were obtained and designated pGSBC1 and pGSBC2.…”

“…(iii) Heterologous production of goadsporin in S. lividans 24) The entire set of goadsporin biosynthetic gene cluster was heterologously expressed in S. lividans by using pGSBC1, and the production of goadsporin in the surrogate host was GodA is translated by ribosomes, and GodB and GodC deliver GodA to the cell membrane. (2) GodD, GodE, GodF, and GodG catalyze the post-translational modification of GodA (hetero cyclization and dehydration).…”