DIFFERENT SITES OF ACIVICIN BINDING AND INACTIVATION OF GAMMA-GLUTAMYL TRANSPEPTIDASES
DIFFERENT SITES OF ACIVICIN BINDING AND INACTIVATION OF GAMMA-GLUTAMYL TRANSPEPTIDASES
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DOI：
10.1073/pnas.92.6.2360
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发表时间：
1995-03-14
影响因子：
11.1
通讯作者：
MEISTER, A
中科院分区：
文献类型：
--
作者：
SMITH, TK;IKEDA, Y;MEISTER, A
Acivicin是γ -谷氨酰转肽酶（EC 2.3.2.2）的有效抑制剂，该酶在谷胱甘肽代谢中起重要作用。-谷氨酰底物和类似物（如丝氨酸加硼酸盐）阻止了Acivicin的抑制和结合，这与先前的假设一致，即Acivicin和底物结合到相同的酶位点。acivicin使大鼠肾转肽酶失活，导致其以酯形式与Thr-523结合。用Ala-523代替Thr-523的猪酶，在Ser-405位点发生酯化反应，被acivicin抑制。人酶有Thr-524（对应大鼠的Thr-523）；它的失活导致Ser-406的酯化（对应于大鼠和猪的Ser-405）。羟胺处理的阿霉素灭活酶恢复活性和释放阿霉素衍生的三- β -羟谷氨酸部分。结果表明，大鼠酶的活性位点区与人、猪的活性位点区存在显著的结构差异。在人类突变酶中，Thr-524和Ser-406分别或一起被Ala取代，具有酶活性，表明这些氨基酸残基不需要催化。然而，这些残基（以及活性位点附近的另一个残基）的酯化反应有效地阻断了活性位点或阻碍了其功能。Acivicin可以结合在谷氨酰化发生的附近的酶位点；它可以在后一个位点结合，然后被酯交换到另一个酶位点。
Acivicin is a potent inhibitor of gamma-glutamyl transpeptidase (EC 2.3.2.2), an enzyme of importance in glutathione metabolism. Acivicin inhibition and binding are prevented by gamma-glutamyl substrates and analogs (e.g., serine plus borate), consistent with the previous postulate that acivicin and substrates bind to the same enzyme site. Inactivation of rat kidney transpeptidase by acivicin leads to its binding as an ester to Thr-523. The pig enzyme, which has Ala-523 in place of Thr-523, is inhibited by acivicin with esterification at Ser-405. The human enzyme has Thr-524 (corresponding to Thr-523 in rat); its inactivation leads to esterification of Ser-406 (corresponding to Ser-405 in rat and pig). Hydroxylamine treatment of the acivicin-inactivated enzymes restores activity and releases the acivicin-derived threo-beta-hydroxyglutamate moiety. The findings indicate that there are significant structural differences between the active site region of the rat enzyme and the active site regions of the human and pig. Human mutant enzymes in which Thr-524 and Ser-406 were replaced by Ala, separately and together, are enzymatically active, indicating that these amino acid residues are not required for catalysis. However, esterification of these residues (and of another near the active site) effectively blocks the active site or hinders its function. Acivicin can bind at enzyme sites that are close to that at which gamma-glutamylation occurs; it may bind at the latter site and then be transesterified to another enzyme site.