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J Biol Chem. 2017 Apr 28;292(17):7066-7076. doi: 10.1074/jbc.M116.771634. Epub 2017 Mar 10.

Effects of a detergent micelle environment on P-glycoprotein (ABCB1)-ligand interactions.

Author information

1
From the Laboratory of Cell Biology, Center for Cancer Research, NCI, National Institutes of Health, Bethesda, Maryland 20892.
2
From the Laboratory of Cell Biology, Center for Cancer Research, NCI, National Institutes of Health, Bethesda, Maryland 20892 ambudkar@helix.nih.gov.

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 IC50 values 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.

KEYWORDS:

ABC transporter; ATP hydrolysis; P-glycoprotein; detergent micelles; docking; drug resistance; molecular modeling; multidrug transporter; nanodiscs; proteoliposomes

PMID:
28283574
PMCID:
PMC5409473
DOI:
10.1074/jbc.M116.771634
[Indexed for MEDLINE]
Free PMC Article

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