Characterization of R Factor β-Lactamases by the Acidimetric Method

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Abstract
The properties and regulation of β-lactamases produced by certain R factors derived from ampicillin-resistant strains of Escherichia coli and Klebsiella were characterized. β-Lactamase activity was determined by the acidimetric method with phenol red as indicator. The sensitivity of this assay was increased by employing phosphate buffer at a final concentration of 0.4 m m ; as little as 0.05 unit (1 unit equals the amount of β-lactamase that hydrolyzes 1 μmole of benzyl penicillin per hr at p H 7.6 and 25 C) could be detected. This assay is rapid and convenient and appears to be superior to other methods currently employed to assay R factor β-lactamases. Two classes of β-lactamases were distinguished on the basis of substrate profile, heat inactivation, and K m values. The regulation of most of these R factor β-lactamases, like certain other R factor enzymes, is subject to cyclic adenosine monophosphate-mediated catabolite repression.