The Catalytic Activity and Penicillin Sensitivity in the Liquid and Frozen States of Membrane‐Bound and Detergent‐Solubilised Transpeptidase of Streptomyces R61

The K m, app . values of the membrane‐bound transpeptidase of Streptomyces R61 for the donor Ac 2 ‐ l ‐Lys‐ d ‐Ala‐ d ‐Ala and the acceptor Gly‐Gly are not affected by temperature variations when the reaction mixtures are incubated in liquid suspensions. At – 5°C, the incubation can be carried out either in the liquid or in the frozen state. The enzyme is active in the latter state. In the frozen state, the K m, app . value for the acceptor remains unchanged but there is a 3‐fold increase in the maximum velocity, a 10‐fold decrease of the K m, app . value for the donor and a 10‐fold increase of the benzylpenicillin concentration required to inhibit the enzyme activity by 50% (ID 50 value). Temperatures of −35°C or below are required to completely inhibit the membranebound enzyme in the frozen state. Cetyltrimethylammonium bromide extracts the transpeptidase both from the isolated membranes and, with a much higher yield, from the intact mycelium. The extracted enzyme is not active in the frozen state, requires detergent for activity, has decreased K m, app . values for both donor and acceptor, exhibits the same sensitivity to benzylpenicillin and cephalosporin C as the membrane‐bound transpeptidase (in liquid suspensions) and, like this latter enzyme, has no dd ‐carboxypeptidase activity. The detergent‐extracted transpeptidase penetrates gels of Sephadex‐100 and is not sedimented at 2 × g .