title: Purification and characterization of macrolide 2'-phosphotransferase type II from a strain of Escherichia coli highly resistant to macrolide antibiotics.
abstract: The resistance mechanism of Escherichia coli BM2506 to macrolides was found to be due to inactivation. Inactivated oleandomycin was identified as oleandomycin 2'-phosphate by thin-layer chromatography. A new type of macrolide-phosphorylating enzyme, macrolide 2'-phosphotransferase type II (MPH(2')II), was detected, purified 95-fold and its enzymological properties investigated. MPH(2')II was a constitutive intracellular enzyme which showed high levels of activity with both 14-member-ring and 16-member-ring macrolides. The optimum pH for the inactivation of oleandomycin was 8.2 and the optimum temperature of the reaction was 40 degrees C. Enzyme activity was lost by heat treatment at 60 degrees C for 1 min. The isoelectric point and M(r) of the enzyme were 5.3 and 48,000, respectively. Purine nucleotides, such as ITP, GTP and ATP, were effective as cofactors in the inactivation of macrolides. An inhibitory effect of iodine, EDTA, or divalent cations on MPH(2')II activity was observed.
authors.affiliation: Department of Microbiology, Tokyo College of Pharmacy, Japan.
authors.forename: M
authors.initials: M
authors.lastname: Kono
authors.forename: K
authors.initials: K
authors.lastname: O'Hara
authors.forename: T
authors.initials: T
authors.lastname: Ebisu
mesh_terms: D000900:Anti-Bacterial Agents / Q000037:antagonists & inhibitors / Q000494:pharmacology; D004352:Drug Resistance, Microbial; D004926:Escherichia coli / Q000187:drug effects / Q000201:enzymology* / Q000302:isolation & purification; D006801:Humans; D008826:Microbial Sensitivity Tests; D009827:Oleandomycin / Q000037:antagonists & inhibitors / Q000494:pharmacology; D010770:Phosphotransferases / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D017853:Phosphotransferases (Alcohol Group Acceptor); D013379:Substrate Specificity
chemical_list: D000900:Anti-Bacterial Agents; D010770:Phosphotransferases; D017853:Phosphotransferases (Alcohol Group Acceptor); C061369:macrolide 2'-kinase; D009827:Oleandomycin