Kozarich J W, Strominger J L
J Biol Chem. 1978 Feb 25;253(4):1272-8.
Staphylococcus aureus H membranes were found to contain four major binding components: Mr = 115,000; Mr = 100,000 doublet; and Mr = 46,000. The low molecular weight protein bound penicillin reversibly and was purified by prebinding membranes with penicillin prior to affinity chromatography. The purified protein catalyzed transpeptidase and carboxypeptidase reactions using di[14C]acetyl-L-lysyl-D-alanyl-D-alanine as the substrate and glycine and hydroxylamine as the acceptors. In addition, the enzyme catalyzed a penicillinase reaction. Kinetic analysis of these reactions revealed similar Vmax values suggesting that, if there is a single active site, the rate-determining steps (i.e. deacetylation) are similar. Rapid denaturation of the enzyme.substrate complex resulted in the detection of covalent penicilloyl- and diacetyl-L-lysyl-D-alanyl.enzyme complexes by sodium dodecyl sulfate-polyacrylamide gel electrophoresis.
发现金黄色葡萄球菌H膜含有四种主要结合成分：分子量为115,000；分子量为100,000的双峰；以及分子量为46,000。低分子量蛋白质可逆地结合青霉素，并在亲和层析之前通过用青霉素预结合膜来进行纯化。纯化后的蛋白质以二[14C]乙酰-L-赖氨酰-D-丙氨酰-D-丙氨酸作为底物、甘氨酸和羟胺作为受体催化转肽酶和羧肽酶反应。此外，该酶还催化青霉素酶反应。对这些反应的动力学分析显示出相似的Vmax值，这表明，如果存在单一活性位点，那么限速步骤（即脱乙酰化）是相似的。酶-底物复合物的快速变性导致通过十二烷基硫酸钠-聚丙烯酰胺凝胶电泳检测到共价青霉素酰-和二乙酰-L-赖氨酰-D-丙氨酰-酶复合物。
