Ruetz S, Gros P
Department of Biochemistry, McGill University, Montreal, Quebec, Canada.
Cell. 1994 Jul 1;77(7):1071-81. doi: 10.1016/0092-8674(94)90446-4.
P-glycoproteins (P-gps) encoded by the mouse mdr2 and mdr3 genes were expressed in secretory vesicles (SVs) from the yeast mutant sec6-4, and their capacity to function as a lipid translocase/flippase was tested. An assay that uses a fluorescent phosphatidylcholine (PC) analog was developed to quantitate asymmetric lipid distribution in the outer and inner leaflets of the lipid bilayer of these vesicles. Mdr2 expression in SVs caused a time- and temperature-dependent enhancement of PC translocation to the inner leaflet of the membrane. The Mdr2-mediated effect was specific since expression of Mdr3 in these vesicles was without effect on the membrane distribution of PC. Increased Mdr2-mediated PC translocation was strictly ATP and Mg2+ dependent, was abrogated by the ATPase inhibitor vanadate and the P-gp modulator verapamil, but was insensitive to the presence of excess of the multidrug resistance drugs colchicine and vinblastine.
由小鼠mdr2和mdr3基因编码的P-糖蛋白（P-gps）在酵母突变体sec6-4的分泌囊泡（SVs）中表达，并测试了它们作为脂质转位酶/翻转酶的功能。开发了一种使用荧光磷脂酰胆碱（PC）类似物的测定法，以定量这些囊泡脂质双层外叶和内叶中不对称脂质分布。SVs中Mdr2的表达导致PC向内膜小叶转位的时间和温度依赖性增强。Mdr2介导的作用是特异性的，因为这些囊泡中Mdr3的表达对PC的膜分布没有影响。Mdr2介导的PC转位增加严格依赖于ATP和Mg2+，被ATP酶抑制剂钒酸盐和P-gp调节剂维拉帕米消除，但对过量的多药耐药药物秋水仙碱和长春碱不敏感。
