Two glycosyltransferases and a glycosidase are involved in oleandomycin modification during its biosynthesis by Streptomyces antibioticus

Abstract

SummaryA 5.2 kb region from the oleandomycin gene cluster in Streptomyces antibioticus located between the oleandomycin polyketide synthase gene and sugar biosynthetic genes was cloned. Sequence analysis revealed the presence of three open reading frames (designated oleI, oleN2 and oleR). The oleI gene product resembled glycosyltransferases involved in macrolide inactivation including the oleD product, a previously described glycosyltransferase from S. antibioticus. The oleN2 gene product showed similarities with different aminotransferases involved in the biosynthesis of 6-deoxyhexoses. The oleR gene product was similar to several glucosidases from different origins. The oleI, oleR and oleD genes were expressed in Streptomyces lividans. OleI and OleD intracellular proteins were partially purified by affinity chromatography in an UDP-glucuronic acid agarose column and OleR was detected as a major band from the culture supernatant. OleI and OleD showed oleandomycin glycosylating activity but they differ in the pattern of substrate specificity: OleI being much more specific for oleandomycin. OleR showed glycosidase activity converting glycosylated oleandomycin into active oleandomycin. A model is proposed integrating these and previously reported results for intracellular inactivation, secretion and extracellular reactivation of oleandomycin.

Dispensable ribosomal resistance to spiramycin conferred by srmA in the spiramycin producer Streptomyces ambofaciens The EMBL/GenBank accession number for the nucleotide sequence described in this paper is AJ223970.

“…Among macrolide producers, pairs of resistance genes have been isolated from the carbomycin producer Streptomyces thermotolerans (Epp et al, 1987), and from ' Micromonospora griseorubida ', the mycinamicin producer (Inouye et al, 1994). Four tylosin-resistance genes have been cloned from the tylosin producer Streptomyces fradiae (Birmingham et al, 1986 ;Baltz & Seno, 1988 ;Rosteck et al, 1991 ;Zalacain & Cundliffe, 1991), and at least five genes are supposed to be involved in oleandomycin resistance in Streptomyces antibioticus (Hernandez et al, 1993 ;Rodriguez et al, 1993 ;Olano et al, 1995 ;Quiros et al, 1998). In contrast, only single resistance genes have been isolated from ' Streptomyces mycarofaciens ', the midecamycin producer (Hara & Hutchinson, 1990), and from the erythromycin producer Sacch.…”

Two new tailoring enzymes, a glycosyltransferase and an oxygenase, involved in biosynthesis of the angucycline antibiotic urdamycin A in Streptomyces fradiae Tü2717 This paper is dedicated to Professor Heinz Floss, a pioneer in the field of antibiotics, on the occasion of his 65th birthday. The GenBank accession numbers for the sequences reported in this paper are AF164960 and AF164961.

“…urdGT2 on a 1n9 kb SalI fragment was ligated into the expression vector pEM4 (Quiros et al, 1998) to create pBF-EM4-1. After transformation of BF-1-1 with this plasmid several colonies were obtained and grown at 28 mC in liquid medium containing thiostrepton.…”

Biosynthesis of elloramycin in Streptomyces olivaceus requires glycosylation by enzymes encoded outside the aglycon cluster

“…Production, transformation, regeneration of protoplasts and conjugation experiments were performed following standard procedures (Kieser et al, 2000). Plasmids pOJ260 (Bierman et al, 1992) and pEM4 (Quiró s et al, 1998) were used for gene inactivation and gene expression, respectively. pCR-Blunt (Invitrogen) was used for cloning of PCR products.…”