Dissection of drug-binding-induced conformational changes in P-glycoprotein.
Dissection of drug-binding-induced conformational changes in P-glycoprotein.
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药物结合诱导的 P-糖蛋白构象变化的剖析。
DOI：
10.1046/j.1432-1327.1998.2550383.x
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发表时间：
1998
期刊：
影响因子：
--
通讯作者：
Zhang,JT
中科院分区：
文献类型：
--
作者：
Wang,G;Pincheira,R;Zhang,JT
P-糖蛋白（Pgp）是一种膜转运ATP酶。它利用ATP水解产生的能量将多效性细胞毒性药物从细胞内运输到细胞外。因此，癌细胞中Pgp表达升高导致多药耐药。现在已知Pgp的构象状态在其催化循环期间发生变化。然而，ATP水解如何与Pgp的药物结合有关尚待确定。在这项研究中，我们使用有限的胰蛋白酶消化Pgp在隔离的内翻膜囊泡，以研究药物对Pgp构象的影响，并确定药物结合的构象状态的Pgp在催化循环。我们发现（a）单独与长春碱或维拉帕米结合可引起Pgp构象变化，但与核苷酸结合引起的变化不同;（B）Pgp底物可能至少有两个结合位点，一个与长春碱和维拉帕米等药物结合，另一个与秋水仙碱和阿霉素等药物结合，（c）ATP和长春碱结合的Pgp构象与单独结合的Pgp构象不同，（d）ADP结合的Pgp不结合长春碱。基于这些观察和我们以前的研究，我们提出了一个模型的药物结合和运输的催化循环的Pgp。
P‐glycoprotein (Pgp) is a membrane‐transport ATPase. It uses energy from ATP hydrolysis to transport pleiotropic cytotoxic drugs from inside to outside of cells. Thus, elevated expression of Pgp in cancer cells causes multidrug resistance. It is now known that the conformational state of Pgp changes during its catalytic cycle. However, how ATP hydrolysis relates to drug binding by Pgp is yet to be determined. In this study, we used limited trypsin digestion of Pgp in isolated inside‐out membrane vesicles to investigate the effects of drugs on Pgp conformation and to determine the drug‐bound conformational states of Pgp in the catalytic cycle. We found that (a) binding of vinblastine or verapamil alone can cause a conformational change in Pgp, but the change induced by the drug binding is different from that induced by nucleotide binding, (b) there may be at least two binding sites for Pgp substrates, one for drugs such as vinblastine and verapamil and the other for drugs such as colchicine and adriamycin, (c) the conformation of Pgp bound by ATP and vinblastine is different from the conformation bound by either one alone, and (d) the ADP‐bound Pgp does not bind vinblastine. Based on these observations and our previous studies, we propose a model for drug binding and transport in the catalytic cycle of Pgp.