Drug-stimulated nucleotide trapping in the human multidrug transporter MDR1. Cooperation of the nucleotide binding domains

Details

Serval ID
serval:BIB_E72E6CCBCDF9
Type
Article: article from journal or magazin.
Collection
Publications
Institution
Title
Drug-stimulated nucleotide trapping in the human multidrug transporter MDR1. Cooperation of the nucleotide binding domains
Journal
Journal of Biological Chemistry
Author(s)
Szabo  K., Welker  E., Bakos  E., Muller  M., Roninson  I., Varadi  A., Sarkadi  B.
ISSN
0021-9258 (Print)
Publication state
Published
Issued date
04/1998
Volume
273
Number
17
Pages
10132-8
Notes
Journal Article Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S. --- Old month value: Apr 24
Abstract
The human multidrug transporter (MDR1 or P-glycoprotein) is an ATP-dependent cellular drug extrusion pump, and its function involves a drug-stimulated, vanadate-inhibited ATPase activity. In the presence of vanadate and MgATP, a nucleotide (ADP) is trapped in MDR1, which alters the drug binding properties of the protein. Here, we demonstrate that the rate of vanadate-dependent nucleotide trapping by MDR1 is significantly stimulated by the transported drug substrates in a concentration-dependent manner closely resembling the drug stimulation of MDR1-ATPase. Non-MDR1 substrates do not modulate, whereas N-ethylmaleimide, a covalent inhibitor of the ATPase activity, eliminates vanadate-dependent nucleotide trapping. A deletion in MDR1 (Delta amino acids 78-97), which alters the substrate stimulation of its ATPase activity, similarly alters the drug dependence of nucleotide trapping. MDR1 variants with mutations of key lysine residues to methionines in the N-terminal or C-terminal nucleotide binding domains (K433M, K1076M, and K433M/K1076M), which bind but do not hydrolyze ATP, do not show nucleotide trapping either with or without the transported drug substrates. These data indicate that vanadate-dependent nucleotide trapping reflects a drug-stimulated partial reaction of ATP hydrolysis by MDR1, which involves the cooperation of the two nucleotide binding domains. The analysis of this drug-dependent partial reaction may significantly help to characterize the substrate recognition and the ATP-dependent transport mechanism of the MDR1 pump protein.
Keywords
Adenine Nucleotides/*metabolism Adenosine Triphosphatases/antagonists & inhibitors Binding Sites Cyclosporine/pharmacology Enzyme Inhibitors/pharmacology Ethylmaleimide/pharmacology Fluoresceins/pharmacology Fluorouracil/pharmacology Humans P-Glycoprotein/*metabolism Protein Binding Recombinant Proteins/metabolism Verapamil/pharmacology
Pubmed
Web of science
Open Access
Yes
Create date
24/01/2008 14:40
Last modification date
20/08/2019 16:10
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