Carboxy groups as essential residues in β-lactamases

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Abstract
1. .beta.-Lactamases were divided into classes A, B and C on the basis of their amino acid sequences. 2. .beta.-Lactamases were incbuated at pH 4.0 with the carboxy-group reagent 1-(3-dimethylaminopropyl)-3-ethylcarbodi-imide plus a coloured nucleophile and the extents of inactivation and nucleophile incorporation were monitored. 3. Two class A enzymes (from Bacillus cereus and Bacillus licheniformis) and two class C enzymes (from Enterobacter cloacae P99 and Pseudomonas aeruginosa) were examined. 4. All four enzymes were inactivated, with total inactivation corresponding to the incorporation of approx. 2-3 mol of nucleophile/mol of enzyme. 5. In the case of .beta.-lactamaes I from Bacillus cereus, some 53% of the incorporated nucleophile was located on glutamic acid-168 in the amino acid sequence.