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Nucleotide binding is the critical regulator of ABCG2 conformational transitions

Authors :
Zsuzsanna Gyöngy
Gábor Mocsár
Éva Hegedűs
Thomas Stockner
Zsuzsanna Ritter
László Homolya
Anita Schamberger
Tamás I Orbán
Judit Remenyik
Gergely Szakacs
Katalin Goda
Source :
eLife, Vol 12 (2023)
Publication Year :
2023
Publisher :
eLife Sciences Publications Ltd, 2023.

Abstract

ABCG2 is an exporter-type ABC protein that can expel numerous chemically unrelated xeno- and endobiotics from cells. When expressed in tumor cells or tumor stem cells, ABCG2 confers multidrug resistance, contributing to the failure of chemotherapy. Molecular details orchestrating substrate translocation and ATP hydrolysis remain elusive. Here, we present methods to concomitantly investigate substrate and nucleotide binding by ABCG2 in cells. Using the conformation-sensitive antibody 5D3, we show that the switch from the inward-facing (IF) to the outward-facing (OF) conformation of ABCG2 is induced by nucleotide binding. IF-OF transition is facilitated by substrates, and hindered by the inhibitor Ko143. Direct measurements of 5D3 and substrate binding to ABCG2 indicate that the high-to-low affinity switch of the drug binding site coincides with the transition from the IF to the OF conformation. Low substrate binding persists in the post-hydrolysis state, supporting that dissociation of the ATP hydrolysis products is required to reset the high substrate affinity IF conformation of ABCG2.

Details

Language :
English
ISSN :
2050084X
Volume :
12
Database :
Directory of Open Access Journals
Journal :
eLife
Publication Type :
Academic Journal
Accession number :
edsdoj.20c8b08529401da60b87c02b59cc6f
Document Type :
article
Full Text :
https://doi.org/10.7554/eLife.83976