Transmembrane helix 12 modulates progression of the ATP catalytic cycle in ABCB1

Crowley, Emily, O'Mara, Megan L., Reynolds, Catherine, Tieleman, D. Peter, Storm, Janet, Kerr, Ian D. and Callaghan, Richard (2009) Transmembrane helix 12 modulates progression of the ATP catalytic cycle in ABCB1. Biochemistry, 48 26: 6249-6258. doi:10.1021/bi900373x


Author Crowley, Emily
O'Mara, Megan L.
Reynolds, Catherine
Tieleman, D. Peter
Storm, Janet
Kerr, Ian D.
Callaghan, Richard
Title Transmembrane helix 12 modulates progression of the ATP catalytic cycle in ABCB1
Journal name Biochemistry   Check publisher's open access policy
ISSN 0006-2960
Publication date 2009-05-20
Year available 2009
Sub-type Article (original research)
DOI 10.1021/bi900373x
Open Access Status DOI
Volume 48
Issue 26
Start page 6249
End page 6258
Total pages 10
Editor Richard N Armstrong
Place of publication United States
Publisher American Chemical Society
Language eng
Subject C1
030403 Characterisation of Biological Macromolecules
970111 Expanding Knowledge in the Medical and Health Sciences
Abstract Understanding the process that underlies multidrug recognition and efflux by P-glycoprotein (ABCB1) remains a key biological challenge. Structural data have recently become available for the murine and Caenorhabditis elegans homologues of ABCB1; however all structures were obtained in the absence of nucleotide. A feature of these structures was the presence of a central cavity that is inaccessible from the extracellular face of the protein. To determine the conformational dynamics of this region several residues in transmembrane helices TM6 (331, 343 and 354) and TM12 (980) were mutated to cysteine. Based upon structural predictions, these residues are proposed to line, or reside proximal to, the central cavity. The mutant isoforms were labelled with a paramagnetic probe enabling the application of EPR spectroscopic methods. Power saturation EPR spectra were recorded in the presence of hydrophobic (O2 ) or hydrophilic (NiEDDA) quenching agents to study the local environment of each residue. ABCB1 was trapped in both its nucleotide-bound and post-hydrolytic conformations and EPR spectra were again recorded in the presence and absence of quenching agents. The EPR line shapes provide information on the movements of these residues within TM6 and TM12 during ATP hydrolysis. Rationalization of the data with molecular dynamic simulations indicates that the cavity is converted to a configuration open to the aqueous phase following nucleotide binding, thereby suggesting alternating access to the cavity on opposite sides of the membrane during translocation.
Keyword ABC transporter
P-glycoprotein
cancer chemotherapy
membrane transport
multidrug resistance
Q-Index Code C1
Q-Index Status Confirmed Code
Grant ID 085768
094392
094392/B/10/Z
Institutional Status UQ

Document type: Journal Article
Sub-type: Article (original research)
Collections: 2010 Higher Education Research Data Collection
School of Chemistry and Molecular Biosciences
 
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Created: Thu, 03 Sep 2009, 17:48:35 EST by Mr Andrew Martlew on behalf of School of Chemistry & Molecular Biosciences