Kayoko S. Tsuchiya

DNA Sequencing and Transcriptional Analysis of the Kasugamycin Biosynthetic Gene Cluster from Streptomyces kasugaensis M338-M1

Streptomyces kasugaensis M338-M1 produces the aminoglycoside antibiotic kasugamycin (KSM). We previously cloned, sequenced and characterized the KSM acetyltransferase, transporter, and some of the biosynthetic genes from this strain. To identify other potential genes in a chromosome walk experiment, a 6.8-kb EcoRI-PstI region immediately downstream from the KSM transporter genes was sequenced. Five open reading frames (designated as kasN, kasO, kasP, kasQ, kasR) and the 5′ region of kasA were found in this region. The genes are apparently co-transcribed as bicistrons, all of which are co-directional except for the kasPQ transcript. Homology analysis of the deduced products of kasN, kasP, kasQ and kasR revealed similarities with known enzymes: KasN, D-amino acid oxidase from Pseudomonas aeruginosa (35% identity); KasP, F420-dependent H4MPT reductase from Streptomyces lavendulae (33% identity); KasQ, UDP-N-acetylglucosamine 2-epimerase from Streptomyces verticillus (45% identity); and KasR, NDP-hexose 3,4-dehydratase from Streptomyces cyanogenus (38% identity); respectively. A gel retardation assay showed that KasT, a putative pathway-specific regulator for this gene cluster, bound to the upstream region of kasN and to the intergenic region of kasQ-kasR, suggesting that the expression of these operons is under the control of the regulator protein.