2 GLYCOSYLTRANSFERASES AND A GLYCOSIDASE ARE INVOLVED IN OLEANDOMYCINMODIFICATION DURING ITS BIOSYNTHESIS BY STREPTOMYCES-ANTIBIOTICUS

A 5.2 kb region from the oleandomycin gene cluster in Streptomyces ant ibioticus located between the oleandomycin polyketide synthase gene an d sugar biosynthetic genes was cloned. Sequence analysis revealed the presence of three open reading frames (designated olel, oleN2 and oleR ). The olel gene product resembled glycosyltransferases involved in ma crolide inactivation including the oleD product, a previously describe d glycosyltransferase from S. antibioticus. The oleN2 gene product sho wed similarities with different aminotransferases involved in the bios ynthesis of 6-deoxyhexoses. The oleR gene product was similar to sever al glucosidases from different origins. The olel, oleR and oleD genes were expressed in Streptomyces lividans. Olel and OleD intracellular p roteins were partially purified by affinity chromatography in an UDP-g lucuronic acid agarose column and OleR was detected as a major band fr om the culture supernatant. Olel and OleD showed oleandomycin glycosyl ating activity but they differ in the pattern of substrate specificity : Olel being much more specific for oleandomycin. OleR showed glycosid ase activity converting glycosylated oleandomycin into active oleandom ycin. A model is proposed integrating these and previously reported re sults for intracellular inactivation, secretion and extracellular reac tivation of oleandomycin.