CHARACTERIZATION AND FUNCTIONAL RECONSTITUTION OF THE MULTIDRUG TRANSPORTER

被引:48
作者
SHAROM, FJ
机构
[1] Guelph-Waterloo Centre for Graduate Work in Chemistry, Department of Chemistry and Biochemistry, University of Guelph, Guelph, NIG 2W1, Ontario
关键词
MULTIDRUG RESISTANCE; MULTIDRUG TRANSPORTER; P-GLYCOPROTEIN; DETERGENT EXTRACTION; ATPASE; INHIBITORS; ACTIVE TRANSPORT; COLCHICINE; RECONSTITUTION; PROTEOLIPOSOMES;
D O I
10.1007/BF02110326
中图分类号
Q6 [生物物理学];
学科分类号
071011 ;
摘要
P-Glycoprotein, the multidrug transporter, is isolated from the plasma membrane of CH(R)C5 cells using a selective two-step detergent extraction procedure. The partially purified protein displays a high level of ATPase activity, which has a high K-M for ATP, is stimulated by drugs, and can be distinguished from that of other membrane ATPases by its unique inhibition profile. Delipidation completely inactivates ATPase activity, which is restored by the addition of fluid lipid mixtures. P-Glycoprotein was reconstituted into lipid bilayers with retention of both drug transport and ATPase activity. Proteoliposomes containing P-glycoprotein display osmotically sensitive ATP-dependent accumulation of H-3-colchicine in the vesicle lumen. Drug transport is active, generating a stable 5.6-fold concentration gradient, and can be blocked by compounds in the multidrug resistance spectrum. Reconstituted P-glycoprotein also exhibits a high level of ATPase activity which is further stimulated by various drugs. P-Glycoprotein therefore functions as an active drug transporter with constitutive ATPase activity.
引用
收藏
页码:15 / 22
页数:8
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