Accumulation of the angucycline antibiotic rabelomycin after disruption of an oxygenase gene in the jadomycin B biosynthetic gene cluster of Streptomyces venezuelae

• Published in: Microbiology (Society for General Microbiology) Vol. 142; no. 1; pp. 123 - 132
• Main Authors: Yang, Keqian, Han, Lei, Ayer, Stephen W, Vining, Leo C
• Format: Journal Article
• Language: English
• Published: Reading Soc General Microbiol 01.01.1996; Society for General Microbiology
• Subjects: Amino Acid Sequence; Anthraquinones - classification; Anthraquinones - metabolism; Anti-Bacterial Agents - classification; Anti-Bacterial Agents - metabolism; Bacteriology; Base Sequence; Biological and medical sciences; Biology of microorganisms of confirmed or potential industrial interest; Biotechnology; Fundamental and applied biological sciences. Psychology; Genes, Bacterial; Genetics; Isoquinolines - metabolism; Microbiology; Mission oriented research; Models, Biological; Molecular Sequence Data; Multienzyme Complexes - genetics; Multigene Family; Mutagenesis; Open Reading Frames; Oxygenases - deficiency; Oxygenases - genetics; Restriction Mapping; Sequence Analysis, DNA; Sequence Homology, Amino Acid; Streptomyces - enzymology; Streptomyces - genetics; Gene cluster; Streptomycetaceae; Nucleotide sequence; Streptomyces venezuelae; Sequence alignment; Biosynthesis; Metabolic pathway; Actinomycetales; Antibiotic; Bacteria; Actinomycetes; Comparative study; Aminoacid sequence; Structural analysis
• ISSN: 1350-0872; 1465-2080
• DOI: 10.1099/13500872-142-1-123

3 Author for correspondence: Leo C. Vining. Tel. +1 902 494 2040. Fax. +1 902 494 3736. e-mail: lvining@ac.dal.ca 1 Department of Biology, Dalhousie University, Halifax, Nova Scotia, Canada B3H 4J1 2 Institute for Marine Biosciences, National Research Council of Canada, Halifax, Nova Scotia, Canada B3H 3Z1 ABSTRACT DNA from a region downstream of and overlapping the polyketide synthase (PKS) gene cluster for jadomycin B biosynthesis in Streptomyces venezuelae was cloned and sequenced. Analysis of the nucleotide sequence located one complete ORF (ORF6), an incomplete one representing the 3' region of ORF4 in the PKS cluster, and a second incomplete one (ORF7). The deduced amino acid sequences for ORFs 6 and 7 resemble those of oxygenases. Since a plausible biosynthetic pathway for jadomycin B includes an angular polyketide intermediate that undergoes oxidative ring fission before condensation with an amino acid, we subcloned one of the presumptive oxygenase genes (ORF6) in a segregationally unstable shuttle vector (pHJL400) and disrupted it by inserting the gene for apramycin resistance. Transformation of S. venezuelae with the disruption vector and selection for apramycin resistance gave mutants blocked in jadomycin biosynthesis. Southern hybridization confirmed that gene replacement had occurred. Cultures of the mutants accumulated a metabolite identified by comparison with an authentic sample as rabelomycin, a non-nitrogenous polyketide-derived antibiotic originally isolated from Streptomyces olivaceus. Keywords: Streptomyces venezuelae, , angucycline, gene disruption, jadomycin B, polyketide antibiotic Present address: Department of Pharmaceutical Sciences, School of Pharmacy, University of Maryland at Baltimore, 20 North Pine Street, Baltimore, MD 21201, USA.