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. 1970 Jan;101(1):232–239. doi: 10.1128/jb.101.1.232-239.1970

Resistance of Escherichia coli to Penicillins VII. Purification and Characterization of a Penicillinase Mediated by the R Factor R1

R Christer Lindqvist 1, Kurt Nordström 1
PMCID: PMC250474  PMID: 4904236

Abstract

The penicillinase mediated by the R factor R1 in Escherichia coli has been purified and characterized. The purification procedure contained the following three steps: spheroplast formation, chromatography of the spheroplast supernatant fluid on DEAE cellulose, and preparative polyacrylamide-gel electrophoresis. The protein obtained gave only one band in analytical polyacrylamide-gel electrophoresis. To obtain milligram quantities of the enzyme, gel filtration on Sephadex G75 was run before the last step in the purification. By gel filtration on Sephadex G75, the molecular weight was estimated as 22,000. The pH optimum, tested in universal buffer, was 7.0. The turnover numbers for benzylpenicillin, d-ampicillin, and 6-aminopenicillanic acid were 4.2 × 104, 6.3 × 104, and 2.2 × 104 moles of substrate hydrolyzed per min by 1 mole of enzyme, whereas the Michaelis constants were 100, 160, and 440 μm, respectively. Cephalosporins were much poorer substrates for the R1 penicillinase than were the penicillins. The turnover number for cephalosporin C, cephaloridine, and 7-amino-cephalosporanic acid were 2.4 × 103, 5 × 102, and less than 2 × 102, respectively. These properties show that the R1 penicillinase is quite different from the chromosomally mediated penicillinase of E. coli (11). However, the R1 enzyme resembles another R-factor penicillinase previously purified by Richmond and Datta.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

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