Cloning and characterization of polyketide synthase genes for jadomycin B biosynthesis in Streptomyces venezuelae ISP5230

Abstract

Hybridizing fragments in the genomic DNA of Streptomyces venezuelae lSP5230, which produces the jadomycin group of angucycline antibiotics, were detected by probing with act/ DNA from Streptomyces coelicolor A3(2). The hybridizing regions were isolated from a 16.5 kb insert of S. venezuelae DNA recovered from a genomic library cloned in a A replacement vector. Subcloning and sequencing of a 4 8 kb segment of the insert, containing regions hybridizing to act/// as well as act/, identified five open reading frames (ORFs). The deduced polypeptide products of the ORFs closely resemble in sequence the components of streptomycete t y p e 4 polyketide synthases (PKSs) : the ORFI product corresponds to the ketoacyl synthase, and the ORF2 product to a polypeptide closely related to the ketoacyl synthase and involved in determining chain length; the ORF3 product matches the acyl carrier protein; ORF4 encodes a bifunctional cyclase/dehydrase; and ORF5 encodes a ketoreductase. Integration into the chromosomal DNA of a plasmid containing a segment of the ORF2-ORF4 region severely depressed jadomycin B biosynthesis; since the integrant showed no change in growth or spore pigmentation, the cloned PKS genes are presumed to encode enzymes in the pathway for jadomycin biosynthesis.

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

“…The 2.4 kb fragment isolated from SacI-digested i LH7 DNA (Han et al, 1994) was subcloned in both orientations in the Sac1 site of pBluescript I1 SK( + ). The recombinant phagemids (p JV60/61) were digested with exonuclease I11 and ligated to generate overlapping deletion clones.…”

“…Cloning and sequence analysis of a fragment of S. venepuelae ISP5230 genomic DNA that hybridizes with act genes from Streptomyes coelicolor A3(2) established the presence of a polyketide synthase (PKS) gene cluster in S. venextrelae. Selective loss of jadomycin B production when a plasmid containing part of the cloned sequence integrated into the chromosome indicated that the PKS genes are associated with jadomycin biosynthesis (Han et al, 1994).…”

Isolation and antimicrobial activities of actinobacteria closely associated with liquorice plants Glycyrrhiza glabra L. and Glycyrrhiza inflate BAT. in Xinjiang, China

“…Three and nine KSa sequences from G. glabra and G. inflate strains, respectively, formed a cluster with KSa sequences related to the biosynthesis of angucycline antibiotics (Han et al, 1994) (Fig. 3).…”

Functional Analyses of Oxygenases in Jadomycin Biosynthesis and Identification of JadH as a Bifunctional Oxygenase/Dehydrase

“…The atypical angucycline antibiotic jadomycin B (1) and its aglycone jadomycin A (2) are produced in cultures of Streptomyces venezuelae ISP5230 grown under stress conditions in a medium containing isoleucine (6,7). The angucycline origin of jadomycins was verified by accumulation of a well known antibiotic rabelomycin (3) in the jadF disruption mutant VS655 (7) and by production of UWM6 (4) in Streptomyces lividans transformed with the jad PKS gene cluster (i.e.…”