The drug-binding pocket of the human multidrug resistance P-glycoprotein is accessible to the aqueous medium
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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Antimicrobial Resistance (ASM)
Antimicrobial resistance poses a significant threat to the continued successful use of antimicrobial agents for the treatment of bacterial infections.
Antimicrobial Resistance
Antimicrobial resistance poses a significant threat to the continued successful use of antimicrobial agents for the treatment of bacterial infections.