title: Effects of a detergent micelle environment on P-glycoprotein (ABCB1)-ligand interactions.
abstract: P-glycoprotein (P-gp) is a multidrug transporter that uses energy from ATP hydrolysis to export many structurally dissimilar hydrophobic and amphipathic compounds, including anticancer drugs from cells. Several structural studies on purified P-gp have been reported, but only limited and sometimes conflicting information is available on ligand interactions with the isolated transporter in a dodecyl-maltoside detergent environment. In this report we compared the biochemical properties of P-gp in native membranes, detergent micelles, and when reconstituted in artificial membranes. We found that the modulators zosuquidar, tariquidar, and elacridar stimulated the ATPase activity of purified human or mouse P-gp in a detergent micelle environment. In contrast, these drugs inhibited ATPase activity in native membranes or in proteoliposomes, with IC50values in the 10-40 nm range. Similarly, a 30-150-fold decrease in the apparent affinity for verapamil and cyclic peptide inhibitor QZ59-SSS was observed in detergent micelles compared with native or artificial membranes. Together, these findings demonstrate that the high-affinity site is inaccessible because of either a conformational change or binding of detergent at the binding site in a detergent micelle environment. The ligands bind to a low-affinity site, resulting in altered modulation of P-gp ATPase activity. We, therefore, recommend studying structural and functional aspects of ligand interactions with purified P-gp and other ATP-binding cassette transporters that transport amphipathic or hydrophobic substrates in a detergent-free native or artificial membrane environment.
authors.affiliation: From the Laboratory of Cell Biology, Center for Cancer Research, NCI, National Institutes of Health, Bethesda, Maryland 20892.
authors.forename: Suneet
authors.initials: S
authors.lastname: Shukla
authors.affiliation: From the Laboratory of Cell Biology, Center for Cancer Research, NCI, National Institutes of Health, Bethesda, Maryland 20892.
authors.forename: Biebele
authors.initials: B
authors.lastname: Abel
authors.affiliation: From the Laboratory of Cell Biology, Center for Cancer Research, NCI, National Institutes of Health, Bethesda, Maryland 20892.
authors.forename: Eduardo E
authors.initials: EE
authors.lastname: Chufan
authors.affiliation: From the Laboratory of Cell Biology, Center for Cancer Research, NCI, National Institutes of Health, Bethesda, Maryland 20892 ambudkar@helix.nih.gov.
authors.forename: Suresh V
authors.initials: SV
authors.lastname: Ambudkar
mesh_terms: D018435:ATP Binding Cassette Transporter, Subfamily B / Q000737:chemistry; D000166:Acridines / Q000737:chemistry; D000255:Adenosine Triphosphate / Q000737:chemistry; D000818:Animals; D016367:Baculoviridae / Q000378:metabolism; D001665:Binding Sites; D003902:Detergents / Q000737:chemistry*; D003986:Dibenzocycloheptenes / Q000737:chemistry; D016503:Drug Delivery Systems; D004353:Drug Evaluation, Preclinical; D005960:Glucosides / Q000737:chemistry; D006801:Humans; D006868:Hydrolysis; D020128:Inhibitory Concentration 50; D007313:Insecta; D008024:Ligands; D051379:Mice; D008823:Micelles; D010456:Peptides, Cyclic / Q000737:chemistry; D011485:Protein Binding; D011804:Quinolines / Q000737:chemistry; D044005:Tetrahydroisoquinolines / Q000737:chemistry; D014700:Verapamil / Q000737:chemistry
chemical_list: C513055:ABCB1 protein, human; D018435:ATP Binding Cassette Transporter, Subfamily B; D000166:Acridines; D003902:Detergents; D003986:Dibenzocycloheptenes; D005960:Glucosides; D008024:Ligands; D008823:Micelles; D010456:Peptides, Cyclic; D011804:Quinolines; D044005:Tetrahydroisoquinolines; C539646:cyclic-tris-(R)-valineselenazole; C040358:dodecyl maltoside; C095179:zosuquidar trihydrochloride; D000255:Adenosine Triphosphate; D014700:Verapamil; C402343:tariquidar; C083501:Elacridar
keywords: ABC transporter; ATP hydrolysis; P-glycoprotein; detergent micelles; docking; drug resistance; molecular modeling; multidrug transporter; nanodiscs; proteoliposomes