Drugs, ionophoric peptides, and steroids as substrates of the yeast multidrug transporter Pdr5p
Drugs, ionophoric peptides, and steroids as substrates of the yeast multidrug transporter Pdr5p
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DOI：
10.1074/jbc.271.49.31543
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发表时间：
1996-12-06
影响因子：
4.8
通讯作者：
Goffeau, A
中科院分区：
文献类型：
--
作者：
Kolaczkowski, M;vanderRest, M;Goffeau, A
Pdr5p是酿酒酵母的三磷酸腺苷结合盒转运体，对几种无关药物产生耐药性。它在Pdr1p转录因子突变体中的高产量使我们能够研究多药转运和底物专一性的分子机制。我们开发了体内和体外检测Pdr5p介导的药物转运的新方法。我们表明，尽管Pdr5p与哺乳动物P-糖蛋白的序列同源性很低，并且拓扑结构颠倒，但Pdr5p与它们有共同的底物。Pdr5p以能量依赖的方式从完整的酵母细胞中分泌罗丹明6G和123。来自Pdr5p高产菌株的质膜制剂显示出依赖于ATP水解的、渗透敏感的罗丹明6G荧光猝灭。许多抗癌药物，如长春新碱、长春新碱、紫杉醇和维拉帕米，以及离子受体多肽和类固醇的微摩尔浓度都竞争性地抑制这种猝灭。相比之下，其他抗癌药物，如秋水仙碱和一些多药耐药调节剂，如奎尼丁，施加非竞争性抑制。我们的实验系统为分析多药物转运蛋白底物和抑制剂的结构-功能关系开辟了新的可能性。
Pdr5p is the yeast Saccharomyces cerevisiae ATP-binding cassette transporter conferring resistance to several unrelated drugs. Its high overproduction in Pdr1p transcription factor mutants allows us to study the molecular mechanism of multidrug transport and substrate specificity. We have developed new in vivo and in vitro assays of Pdr5p-mediated drug transport. We show that in spite of little sequence homology, and inverted topology in respect to that of mammalian P-glycoproteins, Pdr5p shares with them common substrate. Pdr5p extrudes rhodamines 6G and 123, from intact yeast cells in an energy dependent manner. Plasma membrane preparations from a Pdr5p-overproducing strain exhibit ATP hydrolysis-dependent, osmotically sensitive rhodamine 6G fluorescence quenching. The quenching is competitively inhibited by micromolar concentrations of many anticancer drugs, such as vinblastine, vincristine, taxol, and verapamil, and of ionophoric peptides as well as steroids. In contrast, other anticancer drugs, like colchicine and some multidrug resistance modifiers, such as quinidine, exert noncompetitive inhibition. Our experimental system opens new possibilities for the analysis of structure-function relationship of multidrug transporter substrates and inhibitors.