Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P50583 (asymmetrical)
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The exocellular beta-lactamase (penicillin amido-beta-lactamhydrolase, EC 3.5.2.6) of Actinomadura R39 consists of one single polypeptide chain of molecular weight about 15 200. It exhibits a highly asymmetrical shape, has a low isoelectric point (at pH 5.0) and contains about 9.3% (w/w) of a polydeoxyribonucleotide with which it forms a rather stable complex. Removal of a substantial amount of this deoxyribonucleotide by treatment with DNAase I has no effect on the enzyme activity. The beta-lactamase has a wide spectrum of activity. Penicillins and delta 3-cephalosporins can be either good or poor substrates. Oxacillin, which is a poor substrate of most beta-lactamases from Gram-positive bacteria, is a good substrate of the beta-lactamase of Actinomadura R39. Its best substrate, however, is nitrocefin (kcat/Km: 2300 000 M-1.s-1; catalytic centre activity: 210 s-1). The kcat/Km values observed with some penicillins and delta 3-cephalosporins are similar to the values of the bimolecular rate constants that govern the formation of the acyl-enzyme intermediates between these antibiotics and the serine D-alanyl-D-alanine peptidase that is also secreted by the same strain Actinomadura R39. Such a relationship, however, is not observed with all the beta-lactam compounds tested.
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PMID:Purification and properties of the exocellular beta-lactamase of Actinomadura strain R39. 697 97

Carbapenem-hydrolyzing beta-lactamase from Serratia marcescens FHSM4055 was purified 926-fold by means of carboxylmethyl Sephadex C-50, Sephacryl S-200, and Mono S column chromatography. The molecular weight was 30,000 by SDS-PAGE and the isoelectric point was 8.7. The enzyme activity was inhibited by EDTA, and restored by adding zinc (II) or manganese (II). It was inhibited by p-chloromercuribenzoate and iodine as well as the heavy metals, Hg (II), Fe (II), Fe (III), and Cu (II). These results indicate that the enzyme is a metallo-beta-lactamase and that the SH-group of only one cysteine residue probably binds to the metal ion, thus contributing to the stability of the enzyme active center. The specific constant (kcat/Km) showed that the enzyme hydrolyzed various beta-lactam antibiotics such as carbapenems, cephalosporins, moxalactam, cephamycins, and penicillins other than monobactams. Ampicillin and piperacillin with respective amino- and imino-groups, ceftazidime with a carboxypropyloxyimino-group, and cefclidin with a carbamoylquinuclidine-group were poor substrates among the beta-lactam antibiotics other than the monobactams tested. The plots of the turnover number (kcat) against pH for the hydrolysis of cephaloridine gave an asymmetrical curve with the 'tail' on the acid side (pK1, 5.9; pK2, 9.0; pK3, 10.8), whereas those of kcat/Km gave a bell-shaped curve (pK1, 5.8; pK2, 9.8). Both results suggest that two ionic forms of an intermediate yield the same product at different rates and that the enzyme is stable under alkaline conditions.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Purification and characterization of metallo-beta-lactamase from Serratia marcescens. 778 75