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Purification and characterization of penicillinases from Salmonella typhimurium and Escherichia coli

1970; Elsevier BV; Volume: 139; Issue: 2 Linguagem: Inglês

10.1016/0003-9861(70)90479-0

1096-0384

Harold C. Neu, Elaine B. Winshell,

Cancer therapeutics and mechanisms

A penicillinase (penicillin β-lactamase) has been purified from a strain of Salmonella typhimurium in which synthesis of the enzyme is mediated by a resistance-transfer factor. The enzyme is located at the surface of the cell and completely released by the cold osmotic shock technique. The penicillinase is not subject to induction by alteration of growth conditions nor is it induced by nonhydrolyzable penicillins. The enzyme can be prepared by DEAE-cellulose and hydroxylapatite column chromatography. The pH optimum is 7.0. The molecular weight is 30,000 with an S20,w of 2.9. Cysteine is not present in the molecule. The penicillinase has a Km of 2 × 10−5 with benzylpenicillin. Semisynthetic substituted penicillins such as methicillin-6-(2,6-dimethyl benzamido) penicillanic acid act as competitive inhibitors of the hydrolysis of benzylpenicillin. Penicillinase resistance has been transferred from Salmonella typhimurium to Escherichia coli, Shigella sonnei, Serralia marcescens, and Proteus mirabilis. The pcnicillinases of all of these except Proteus mirabilis can be released from the cells by osmotic shock. Penicillinases purified from Escherichia coli, Serratia, and Shigella into which the resistance-transfer factor from Salmonella typhimurium had been introduced were similar in molecular weight (30,000), pH optimum (7), Km, and electrophoretic mobility in polyacrylamide. Antiserum prepared against the S. typhimurium penicillinase cross reacts with the penicillinases of these other organisms. Penicillinase purified from an E. coli strain and a S. typhimurium strain which lack a resistance-transfer factor but release penicillinase when subjected to osmotic shock show similar properties, amino acid composition, and antigenicity with the penicillinases from resistance transfer-factor strains of Enterobacteriaceae.