Studies with novel Pdr5p substrates demonstrate a strong size dependence for xenobiotic efflux
Studies with novel Pdr5p substrates demonstrate a strong size dependence for xenobiotic efflux
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DOI：
10.1074/jbc.m210908200
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发表时间：
2003-02-21
影响因子：
4.8
通讯作者：
May, L
中科院分区：
文献类型：
--
作者：
Golin, J;Ambudkar, SV;May, L
酵母(酿酒酵母)多药转运体Pdr5p可外排多种结构和作用方式不同的底物。以前的工作表明，分子大小和电离可能是重要的参数。在这项研究中，我们比较了同基因的pdr5和pdr5菌株对化学结构相似的假定底物的相对敏感性。使用了三个系列：含咪唑的化合物、三烷基氯化锡化合物和四烷基锡化合物。我们证明了Pdr5p转运蛋白能够介导既不电离也不存在电子对供体的底物的运输，并且这些底物的结构比那些由人MDR1编码的P-糖蛋白运输的底物简单得多。此外，底物的大小是关键的，并且与疏水性的任何要求无关。底物的表面体积大于90埃(3)，通过分子模拟确定，在200-225A(3)附近有最佳响应。采用测定[H-3]氯霉素和[H-3]三苯咪唑细胞外排的方法。咪唑类化合物对氯霉素转运的抑制呈浓度依赖性，而有机锡化合物和抗肿瘤药物阿霉素均未观察到。相反，几种有机锡化合物是三苯咪唑外排的有效抑制剂，但Pdr5p底物四丙基锡在这两种检测中都无效。这证明了Pdr5p上至少存在三个与哺乳动物P-糖蛋白行为不同的底物结合位点。证据还表明，一些底物能够在一个以上的位置相互作用。令人惊讶的是，Pdr5p介导了对四烷基锡的抗性，这表明其中一个位点可能只使用疏水相互作用来结合底物。
The yeast (Saccharomyces cerevisiae) multidrug transporter Pdr5p effluxes a broad range of substrates that are variable in structure and mode of action. Previous work suggested that molecular size and ionization could be important parameters. In this study, we compared the relative sensitivity of isogenic PDR5 and pdr5 strains toward putative substrates that are similar in chemical structure. Three series were used: imidazole-containing compounds, trialkyltin chlorides, and tetraalkyltin compounds. We demonstrate that the Pdr5p transporter is capable of mediating transport of substrates that neither ionize nor have electron pair donors and that are much simpler in structure than those transported by the human MDR1-encoded P-glycoprotein. Furthermore, the size of the substrate is critical and independent of any requirement for hydrophobicity. Substrates have surface volumes greater than 90 Angstrom(3) with an optimum response at similar to200-225 A(3) as determined by molecular modeling. Assays measuring the efflux from cells of [H-3]chloramphenicol and [H-3]tritylimidazole were used. A concentration-dependent inhibition of chloramphenicol transport was observed with imidazole derivatives but not with either the organotin compounds or the antitumor agent doxorubicin. In contrast, several of the organotin compounds were potent inhibitors of tritylimidazole efflux, but the Pdr5p substrate tetrapropyltin was ineffective in both assays. This argues for the existence of at least three substrate-binding sites on Pdr5p that differ in behavior from those of the mammalian P-glycoprotein. Evidence also indicates that some substrates are capable of interacting at more than one site. The surprising observation that Pdr5p mediates resistance to tetraalkyltins suggests that one of the sites might use only hydrophobic interactions to bind substrates.