CHARACTERIZATION OF OXA-9, A BETA-LACTAMASE ENCODED BY THE MULTIRESISTANCE TRANSPOSON TN1331

The enzyme OXA-9, an oxacillinase-carbenicillinase, is encoded by the bla(OXA-9) gene which was originally found within the structure of Tn1 331, a multiresistance transposon first isolated from a clinical Klebs iella pneumoniae strain. Studies to characterize OXA-9 demonstrated th at it has a pi of 6.9 and the optimal pH for enzyme activity was betwe en 7.7 and 8.2. When total soluble extracts were preincubated at 37 de grees C and at 42 degrees C, enzyme activity decayed to approximately 56% of the original value after 6 hrs. at 37 degrees C and to 50% afte r 30 min. at 42 degrees C. Enzymatic activity of OXA-9 was inhibited i n the presence of p-chloromercuribenzoate, cloxacillin and clavulanic acid, but not by 200 mM sodium chloride. The inhibition by p-chloromer curibenzoate may indicate the presence of a cysteine residue playing a role in the catalytic activity of the enzyme. The OXA-9 enzyme was re leased by osmotic shock of E. coli cells harboring a recombinant clone including the bla(OXA-9) gene indicating that it is located in the pe riplasmic space of the cells. The OXA-9 enzyme was purified from solub le protein extracts of E. coli cells carrying a recombinant clone incl uding the bla(OXA-9) by ion exchange chromatography.