Two new tailoring enzymes, a glycosyltransferase and an oxygenase, involved in biosynthesis of the angucycline antibiotic urdamycin A in Streptomycesfradiae Tu2717

Urdamycin A, the principal product of Streptomyces fradiae Tu2717, is an an gucycline-type antibiotic and anticancer agent containing C-glycosidically linked D-olivose. To extend knowledge of the biosynthesis of urdamycin A th e authors have cloned further parts of the urdamycin biosynthetic gene clus ter. Three new ORFs (urdK, urdJ and urdO) were identified on a 3.35 kb frag ment, and seven new ORFs (urdL, urdM, urdJ2, urdZ1, urdGT2, urdG and urdH) on an 8.05 kb fragment. The deduced products of these genes show similariti es to transporters (urdJ and urdJ2), regulatory genes (urdK), reductases (u rdO),cyclases (urdL) and deoxysugar biosynthetic genes (urdG, urdH ana urdO ). The product of urdM shows striking sequence similarity to oxygenases (N- terminal sequence) as well as reductases (C-terminal sequence), and the ded uced amino acid sequence of urdGT2 resembles those of glycosyltransferases. To the function of urdM and urdGT2, targeted gene inactivation experiments were performed. The resulting urdM deletion mutant strains accumulated pre dominantly rabelomycin, indicating that UrdM is involved in oxygenation at position 12b of urdamycin A. A mutant in which urdGT2 had been deleted prod uced urdamycin I, urdamycin J and urdamycin K instead of urdamycin A. Urdam ycins I, J and K are tetracyclic angucyclinones lacking a C-C connected deo xysugar moiety. Therefore UrdGT2 must catalyse the earliest glycosyltransfe r step in the urdamycin biosynthetic pathway, the C-glycosyltransfer of one NDP-D-olivose.