Identification of a sugar flexible glycosyltransferase from Streptomyces olivaceus, the producer of the antitumor polyketide elloramycin

Background: Elloramycin is an anthracycline-like antitumor drug related to tetracenomycin C which is produced by Streptomyces olivaceus Tu2353. Struct urally is a tetracyclic aromatic polyketide derived from the condensation o f 10 acetate units. Its chromophoric aglycon is glycosylated with a permeth ylated L-rhamnose moiety at the C-8 hydroxy group. Only limited information is available about the genes involved in the biosynthesis of elloramycin. From a library of chromosomal DNA from S. olivaceus, a cosmid (16F4) was is olated that contains part of the elloramycin gene cluster and when expresse d in Streptomyces lividans resulted in the production of a non-glycosylated intermediate in elloramycin biosynthesis, 8-demethyl-tetracenomycin C (8-D MTC). Results: The expression of cosmid 16F4 in several producers of glycosylated antibiotics has been shown to produce tetracenomycin derivatives containin g different 6-deoxysugars. Different experimental approaches showed that th e glycosyltransferase gene involved in these glycosylation events was locat ed in 16F4. Using degenerated oligoprimers derived from conserved amino aci d sequences in glycosyltransferases, the gene encoding this sugar flexible glycosyltransferase (elmGT) has been identified. After expression of elmGT in Streptomyces albus under the control of the erythromycin resistance prom oter, ermEp, it was shown that elmG can transfer different monosaccharides (both L- and D-sugars) and a disaccharide to 8-DMTC. Formation of a diolivo syl derivative in the mithramycin producer Streptomyces argillaceus was fou nd to require the cooperative action of two mithramycin glycosyltransferase s (MtmGI and MtmGII) responsible for the formation of the diolivosyl disacc haride, which is then transferred by ElmGT to 8-DMTC. Conclusions: The ElmGT glycosyltransferase from S. olivaceus Tu2353 can tra nsfer different sugars into the aglycon 8-DMTC. In addition to its natural sugar substrate L-rhamnose, ElmGT can transfer several L- and D-sugars and also a diolivosyl disaccharide into the aglycon 8-DMTC. ElmGT is an example of sugar flexible glycosyltransferase and can represent an important tool for combinatorial biosynthesis. (C) 2001 Elsevier Science Ltd. All rights r eserved.