Purification and characterization of OXA-23 from Acinetobacter baumannii.
Although the existence of bla(OXA-23) is reported in various parts of the world, the product of bla(OXA-23) gene, OXA-23, has not been purified and its kinetic properties are not known. In this study, OXA-23 of Acinetobacter baumannii isolated from Kocaeli University intensive care unit was characterized after purification using recombinant methods. Preliminary results showed that conventional protein purification methods were not effective for purification of OXA-23. Therefore, OXA-23 was fused to maltose-binding protein of Escherichia coli, the fused protein was expressed and purified to homogeneity. Kinetic properties of the pure protein were then studied with substrates e.g., imipenem, meropenem, cefepime, ceftazidime, ampicilline, piperacillin, penicillin G, and nitrocefin. Also clavulanic acid, tazobactam, and sulbactam concentrations that inhibit 50% of OXA-23 enzyme activity were calculated. Modelling of OXA-23 revealed its ionic surface structure, conformation in the fused form and its topology allowing us to make predictions for OXA-23 substrate specificity.
Department Of Medical Biology/KABI Proteomics Laboratory, Kocaeli University Medical School, Kocaeli, Turkey.
Sinem
S
Torol
Murat
M
Kasap
D040981:Acinetobacter baumannii / Q000201:enzymology*; D019818:Clavulanic Acid / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation / Q000187:drug effects; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D007700:Kinetics; D008958:Models, Molecular; D010397:Penicillanic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D013329:Structure-Activity Relationship; D013379:Substrate Specificity; D013407:Sulbactam / Q000494:pharmacology; D013499:Surface Properties; D000078142:Tazobactam; D065093:beta-Lactamase Inhibitors; D001618:beta-Lactamases / Q000302:isolation & purification* / Q000378:metabolism*
D065093:beta-Lactamase Inhibitors; D019818:Clavulanic Acid; D010397:Penicillanic Acid; C466679:beta-lactamase OXA-23; D001618:beta-Lactamases; D013407:Sulbactam; D000078142:Tazobactam