Bruggemann E P, Currier S J, Gottesman M M, Pastan I
Laboratory of Molecular Biology, National Cancer Institute, Bethesda, Maryland 20892.
J Biol Chem. 1992 Oct 15;267(29):21020-6.
To determine the number of drug binding sites that exist on the multidrug transporter, P-glycoprotein, we used azidopine, a dihydropyridine photoaffinity compound that reverses multidrug resistance and labels P-glycoprotein. Azidopine labels P-glycoprotein in two distinct locations: one labeled site is within the amino half of P-glycoprotein between amino acid residues 198 and 440, and the other site is within the carboxy half of the protein. Vinblastine is a cytotoxic drug that is used in cancer chemotherapy and is a substrate for transport by P-glycoprotein. We found that vinblastine inhibits azidopine labeling to approximately the same extent at each labeled site on P-glycoprotein. Because several studies have shown that amino acid residue 185 of P-glycoprotein plays a critical role in some aspects of drug binding and transport, we also studied the effect that amino acid residue 185 has on azidopine labeling. These studies show that azidopine labels both sites equivalently in both wild-type (G185) and mutant (V185) P-glycoproteins. We conclude from our results that the two halves of P-glycoprotein approach each other to form a single binding site for these drugs.
为了确定多药转运蛋白P-糖蛋白上存在的药物结合位点数量，我们使用了叠氮平，一种能逆转多药耐药性并标记P-糖蛋白的二氢吡啶光亲和化合物。叠氮平在两个不同位置标记P-糖蛋白：一个标记位点在P-糖蛋白的氨基端一半内，位于氨基酸残基198和440之间，另一个位点在该蛋白的羧基端一半内。长春碱是一种用于癌症化疗的细胞毒性药物，是P-糖蛋白转运的底物。我们发现长春碱在P-糖蛋白的每个标记位点上抑制叠氮平标记的程度大致相同。由于多项研究表明P-糖蛋白的氨基酸残基185在药物结合和转运的某些方面起关键作用，我们还研究了氨基酸残基185对叠氮平标记的影响。这些研究表明，叠氮平在野生型（G185）和突变型（V185）P-糖蛋白中对两个位点的标记效果相同。我们从结果中得出结论，P-糖蛋白的两半相互靠近，形成了这些药物的单一结合位点。
