The Peptidoglycan Crosslinking Enzyme System in Streptomyces Strains R61, K15 and rimosus. Kinetic Coefficients Involved in the Interactions of the Membrane‐Bound Transpeptidase with Peptide Substrates and β‐Lactam Antibiotics

Abstract

The transpeptidation reaction performed by the membranes of Streptomyces strain R61 fits the general rate equation for an enzyme-catalyzed bimolecular reaction. The same membranes (E) interact with .beta.-lactams (I) to form inactive penicillin.cntdot.enzyme.cntdot.membrane complexes (EI*) of rather high stability, which subsequently break down (E + I .fwdarw. EI* .fwdarw. E + degradation products). The enzyme is regenerated and the antibiotic is released in the form of an inactive metabolite. With benzylpenicillin, the degradation product is benzylpenicilloic acid. The reaction is heat-labile. The 1st step of the reaction (E + I .fwdarw. EI*) is characterized by a 2nd-order rate constant (kformation in M-1 s-1) and the 2nd step (EI* .fwdarw. E + degradation products) by a 1st-order rate constant (kbreakdown in s-1). The effects in vitro of various .beta.-lactams on the membrane-bound transpeptidase, as expressed by the relevant kformation and kbreakdown values, parallel the effects in vivo of the same antibiotics as expressed by their ability to prevent the germination and growth of conidiospores. The kinetic parameters of the transpeptidase that was solubilized with N-cetyl-N,N,N-trimethylammonium bromide with respect to its interaction with both peptide substrates and .beta.-lactam antibiotics are quantitatively different from those of the membrane-bound enzyme. The solubilized enzyme fragments benzylpenicillin with formation of phenylacetylglycine, a reaction which is similar to that catalyzed by the exocellular R61 enzyme. The membranes of S. rimosus and K15 possess an active classic penicillinase. They were not studied but the kinetic coefficients of the corresponding solubilized transpeptidases were determined and compared with those of the solubilized enzyme from strain R61.