Transmembrane helix 12 plays a pivotal role in coupling energy provision and drug binding in ABCB1

Crowley, Emily, O'Mara, Megan L., Kerr, Ian D. and Callaghan, Richard (2010) Transmembrane helix 12 plays a pivotal role in coupling energy provision and drug binding in ABCB1. Febs Journal, 277 19: 3974-3985. doi:10.1111/j.1742-4658.2010.07789.x

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Author Crowley, Emily
O'Mara, Megan L.
Kerr, Ian D.
Callaghan, Richard
Title Transmembrane helix 12 plays a pivotal role in coupling energy provision and drug binding in ABCB1
Journal name Febs Journal   Check publisher's open access policy
ISSN 1742-464X
Publication date 2010-10
Sub-type Article (original research)
DOI 10.1111/j.1742-4658.2010.07789.x
Volume 277
Issue 19
Start page 3974
End page 3985
Total pages 12
Place of publication Oxford, England
Publisher Blackwell Publishing
Collection year 2011
Language eng
Subject C1
0601 Biochemistry and Cell Biology
1101 Medical Biochemistry and Metabolomics
Abstract Describing the molecular details of the multidrug efflux process of ABCB1, in particular the interdomain communication associated with bioenergetic coupling, continues to prove difficult. A number of investigations to date have implicated transmembrane helix 12 (TM12) in mediating communication between the transmembrane domains and nucleotide-binding domains (NBDs) of ABCB1. The present investigation further addressed the role of TM12 in ABCB1 by characterizing its topography during the multidrug efflux process with the use of cysteine-directed mutagenesis. Cysteines were introduced at various positions along TM12 and assessed for their ability to covalently bind thiol-reactive fluorescent probes with differing physiochemical properties. By analysing each isoform in the basal, ATP-bound and posthydrolytic states, it was possible to determine how the local environment of TM12 alters during the catalytic cycle. Labelling with hydrophobic CM and zwitterionic BM was extensive throughout the helix in the basal, prehydrolytic and posthydrolytic states, suggesting that TM12 is in a predominantly hydrophobic environment. Overall, the carboxy region (intracellular half) of TM12 appeared to be more responsive to changes in the catalytic state of the protein than the amino region (extracellular half). Thus, the carboxy region of TM12 is suggested to be responsive to nucleotide binding and hydrolysis at the NBDs and therefore directly involved in interdomain communication. This data can be reconciled with an atomic-scale model of human ABCB1. Taken together, these results indicate that TM12 plays a key role in the progression of the ATP hydrolytic cycle in ABCB1 and, in particular, in coordinating conformational changes between the NBDs and transmembrane domains.
Keyword ABC transporter
bioenergetic coupling
drug resistance
efflux pumps
Human p-glycoprotein
Catalytic cycle
Q-Index Code C1
Q-Index Status Confirmed Code
Institutional Status UQ

Document type: Journal Article
Sub-type: Article (original research)
Collections: Official 2011 Collection
School of Chemistry and Molecular Biosciences
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Citation counts: TR Web of Science Citation Count  Cited 13 times in Thomson Reuters Web of Science Article | Citations
Scopus Citation Count Cited 13 times in Scopus Article | Citations
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Created: Sun, 17 Oct 2010, 00:12:08 EST