[32P]-labeled ATPase was isolated in a highly purified state from Micrococcus lysodeikticus strain PNB grown in medium supplemented with [32P]orthophosphate. Selective extraction procedures allowed us to determine that at least 25% of the firmly bound label belonged to adenine nucleotides, ATP and ADP being present in equimolar amounts. However, no 32P label was found to be part of phospholipids. This was confirmed by purification of the ATPase from cells fed with [2-3H]glycerol. Using the luciferin-luciferase assay we estimated that ATPase freshly isolated by Sephadex chromatography (specific activity 10-14 micromole substrate transformed x min(-1) x mg protein(-1)) contained 2 moles ATP/mole of enzyme. The ratio fell with the age of enzyme and its purification by gel electrophoresis and this was paralleled by a loss of ATPase activity. The endogenous nucleotides were readily exchanged by added ADP or ATP. This result suggests that the sites for tight binding of adenine nucleotides are equivalent, although ADP seems to have a higher affinity for them. The last properties represent a peculiar characteristic of this bacterial ATPase as compared with other bacterial and organelle energy-transducing proteins.

中文翻译：

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无磷脂纯化微球菌溶血微球菌腺苷三磷酸酶中紧密结合的腺嘌呤核苷酸的存在和作用的证据。

[32P] 标记的 ATP 酶以高度纯化的状态从溶血微球菌 PNB 菌株中分离，该菌株在补充有 [32P] 正磷酸盐的培养基中生长。选择性提取程序使我们能够确定至少 25% 的牢固结合标记属于腺嘌呤核苷酸，ATP 和 ADP 以等摩尔量存在。然而，没有发现 32P 标记是磷脂的一部分。这通过从用 [2-3H] 甘油喂养的细胞中纯化 ATP 酶来证实。使用荧光素-荧光素酶测定，我们估计由 Sephadex 层析（比活性 10-14 微摩尔底物转化 x min(-1) x mg 蛋白 (-1)）新鲜分离的 ATPase 含有 2 摩尔 ATP/摩尔酶。该比率随着酶的老化和凝胶电泳的纯化而下降，这与 ATPase 活性的丧失相平行。内源性核苷酸很容易被添加的 ADP 或 ATP 交换。这一结果表明腺嘌呤核苷酸的紧密结合位点是等效的，尽管 ADP 似乎对它们具有更高的亲和力。与其他细菌和细胞器能量转导蛋白相比，最后一个特性代表了这种细菌 ATP 酶的独特特征。