The drug-binding pocket of the human multidrug resistance P-glycoprotein is accessible to the aqueous medium

Biochemistry
Tip W LooDavid M Clarke

Abstract

P-Glycoprotein (P-gp) is an ATP-dependent drug pump that transports a broad range of compounds out of the cell. Cross-linking studies have shown that the drug-binding pocket is at the interface between the transmembrane (TM) domains and can simultaneously bind two different drug substrates. Here, we determined whether cysteine residues within the drug-binding pocket were accessible to the aqueous medium. Cysteine mutants were tested for their reactivity with the charged thiol-reactive compounds sodium (2-sulfonatoethyl)methanethiosulfonate (MTSES) and [2-(trimethylammonium)ethyl)]methanethiosulfonate (MTSET). Residue Ile-306(TM5) is close to the verapamil-binding site. It was changed to cysteine, reacted with MTSES or MTSET, and assayed for verapamil-stimulated ATPase activity. Reaction of mutant I306C(TM5) with either compound reduced its affinity for verapamil. We confirmed that the reduced affinity for verapamil was indeed due to introduction of a charge at position 306 by demonstrating that similar effects were observed when Ile-306 was replaced with arginine or glutamic acid. Mutant I306R showed a 50-fold reduction in affinity for verapamil and very little change in the affinity for rhodamine B or colchicine. MTSES or MTSE...Continue Reading

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Citations

Jun 7, 2008·Microbiology and Molecular Biology Reviews : MMBR·Amy L DavidsonJue Chen
Jan 12, 2007·Biochemistry and Cell Biology = Biochimie Et Biologie Cellulaire·Frances J Sharom
Jan 23, 2009·BMC Structural Biology·Jean-Paul BeckerMartine Prévost
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Jan 1, 2014·Environmental Toxicology and Chemistry·Anastasia GeorgantzopoulouAlbertinka J Murk
Mar 2, 2005·Biochemical and Biophysical Research Communications·Tip W Loo, David M Clarke
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Dec 14, 2005·Archives of Biochemistry and Biophysics·Gregory TomblineAlan E Senior
Oct 8, 2005·Pharmacology & Therapeutics·Mikihisa TakanoTeruo Murakami
Mar 11, 2008·Archives of Biochemistry and Biophysics·Tip W Loo, David M Clarke
Nov 9, 2004·Biochemical and Biophysical Research Communications·Tip W LooDavid M Clarke
Jul 5, 2016·The Science of the Total Environment·Anastasia GeorgantzopoulouAlberTinka J Murk
Aug 24, 2006·The Journal of Biological Chemistry·Tip W LooDavid M Clarke
Feb 4, 2006·The Journal of Membrane Biology·T W Loo, D M Clarke
Mar 25, 2018·Drug Metabolism and Disposition : the Biological Fate of Chemicals·Daria N Trofimova, Roger G Deeley
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Jul 9, 2009·Chemical Reviews·Paul D W Eckford, Frances J Sharom
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