Cloning and analysis of a halogenase gene of Streptomyces sp. 604F from the Arctic Ocean

Abstract:

Abstract:［Objective］ To clone a halogenase gene from halometabolite-producing Streptomyces sp．604F to facilitate identification of potential halometabolites and its biosynthetic gene cluster.［Methods］We used agar block method to detect the antimicrobial activity of Streptomyces sp． 604F． We further amplified the conserved regions of type I polyketide synthase (PKS I)，type II polyketide synthase (PKS II) and nonribosomal peptide synthetase (NRPS) encoding genes by degenerative PCR． We detected halometabolites in fermentation extracts of Streptomyces sp．604F analyzed by liquid chromatography-time of flight mass spectrometry (LC-Tof MS)．Next，we amplified halogenase gene fragment from Streptomyces sp．604F by using degenerative primers targeting reduced flavin adenine dinucleotide (FADH2)-dependent halogenase genes．Finally，we cloned the full halogenase gene and its flanking sequences through high-efficiency thermal asymmetric interlaced PCR (hiTAIL-PCR).［Results］Streptomyces sp．604F showed promising antifungal activity against Candida albicans ATCC 10231，and its genome contained genes encoding PKS I，PKS II，NRPS and a halogenase with 1443 bp．The halogenase is a new non-tryptophan halogenase，and most closely related to halogenases catalyzing the chlorination of glycopeptides.［Conclusion］Streptomyces sp．604F possessed a new non-tryptophan halogenase，which may be involved in halogenation of glycopeptide-like metabolites．The cloning and analysis of this halogenase have provided guidance for searching target halometabolites，and laid the foundation for obtaining the biosynthetic gene cluster.