title: Biosynthesis of the macrolide oleandomycin by Streptomyces antibioticus. Purification and kinetic characterization of an oleandomycin glucosyltransferase.
abstract: The oleandomycin (OM) producer, Streptomyces antibioticus, possesses a mechanism involving two enzymes for the intracellular inactivation and extracellular reactivation of the antibiotic. Inactivation takes place by transfer of a glucose molecule from a donor (UDP-glucose) to OM, a process catalyzed by an intracellular glucosyltransferase. Glucosyltransferase activity is detectable in cell-free extracts concurrent with biosynthesis of OM. The enzyme has been purified 1,097-fold as a monomer, with a molecular mass of 57.1 kDa by a four-step procedure using three chromatographic columns. The reaction operates via a compulsory-order mechanism. This has been shown by steady-state kinetic studies using either OM or an alternative substrate (rosaramycin) and dead-end inhibitors, and isotopic exchange reactions at equilibrium. OM binds first to the enzyme, followed by UDP-glucose. A ternary complex is thus formed prior to transfer of glucose. UDP is then released, followed by the glycosylated oleandomycin (GS-OM).
authors.affiliation: Departamento de Biología Funcional, Universidad de Oviedo, Spain.
authors.forename: L M
authors.initials: LM
authors.lastname: Quirós
authors.forename: J A
authors.initials: JA
authors.lastname: Salas
mesh_terms: D004917:Erythromycin / Q000494:pharmacology; D005964:Glucosyltransferases / Q000302:isolation & purification* / Q000378:metabolism*; D006031:Glycosylation; D007700:Kinetics; D009827:Oleandomycin / Q000096:biosynthesis*; D013303:Streptomyces antibioticus / Q000201:enzymology / Q000378:metabolism*; D014532:Uridine Diphosphate Glucose / Q000494:pharmacology
chemical_list: D004917:Erythromycin; D005964:Glucosyltransferases; D009827:Oleandomycin; D014532:Uridine Diphosphate Glucose