Format

Send to

Choose Destination
J Biol Chem. 2018 Apr 27;293(17):6297-6307. doi: 10.1074/jbc.RA118.002190. Epub 2018 Mar 6.

Conformational dynamics of P-glycoprotein in lipid nanodiscs and detergent micelles reveal complex motions on a wide time scale.

Author information

1
From the Department of Medicinal Chemistry, University of Washington, Seattle, Washington 98195-7610.
2
From the Department of Medicinal Chemistry, University of Washington, Seattle, Washington 98195-7610 winky@uw.edu.

Abstract

P-glycoprotein (P-gp) is a highly substrate-promiscuous efflux transporter that plays a critical role in drug disposition. P-gp utilizes ATP hydrolysis by nucleotide-binding domains (NBDs) to drive transitions between inward-facing (IF) conformations that bind drugs and outward-facing (OF) conformations that release them to the extracellular solution. However, the details of the protein dynamics within either macroscopic IF or OF conformation remain uncharacterized, and the functional role of local dynamics has not been determined. In this work we measured the local dynamics of the IF state of P-gp in lipid nanodiscs and in detergent solution by hydrogen-deuterium (H/D) exchange MS. We observed "EX1 exchange kinetics," or bimodal kinetics, for several peptides distributed in both NBDs, particularly for P-gp in the lipid nanodiscs. Remarkably, the EX1 kinetics occurred on several time scales, ranging from seconds to hours, suggesting highly complex, and correlated, motions. The results indicate at least three distinct conformational states in the ligand-free P-gp and suggest a rough conformational landscape. Addition of excess ATP and vanadate, to favor the OF conformations, caused a generalized, but modest, decrease in H/D exchange throughout the NBDs and slowed the EX1 kinetic transitions of several peptides. The functional implications of the results are consistent with the possibility that conformational selection provides a source of substrate promiscuity.

KEYWORDS:

ATPase; conformational change; conformational selection; drug transport; multidrug resistance; protein conformation; protein dynamic

PMID:
29511086
PMCID:
PMC5925813
DOI:
10.1074/jbc.RA118.002190
[Indexed for MEDLINE]
Free PMC Article

Supplemental Content

Full text links

Icon for HighWire Icon for PubMed Central
Loading ...
Support Center