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Cytosolic region of TM6 in P-glycoprotein: Topographical analysis and functional perturbation by site directed labeling

Storm, Janet; Modok, Szabolcs; O'Mara, Megan; Tieleman, D. Peter; Kerr, Ian; Callaghan, Richard

Description

Reduced intracellular drug accumulation due to the activity of the drug efflux pump ABCB1 is a major mechanism in the resistance of cancer cells to chemotherapy. ABCB1 is a poly specific transporter, and the molecular mechanism of its complex translocation process remains to be elucidated. To understand the process will require information on the regions involved in drug binding and those that couple this event to nucleotide hydrolysis. The present investigation focuses on the cytosolic region...[Show more]

dc.contributor.authorStorm, Janet
dc.contributor.authorModok, Szabolcs
dc.contributor.authorO'Mara, Megan
dc.contributor.authorTieleman, D. Peter
dc.contributor.authorKerr, Ian
dc.contributor.authorCallaghan, Richard
dc.date.accessioned2015-12-13T22:17:28Z
dc.identifier.issn0006-2960
dc.identifier.urihttp://hdl.handle.net/1885/71142
dc.description.abstractReduced intracellular drug accumulation due to the activity of the drug efflux pump ABCB1 is a major mechanism in the resistance of cancer cells to chemotherapy. ABCB1 is a poly specific transporter, and the molecular mechanism of its complex translocation process remains to be elucidated. To understand the process will require information on the regions involved in drug binding and those that couple this event to nucleotide hydrolysis. The present investigation focuses on the cytosolic region of transmembrane helix 6 (TM6), which has been widely attributed with a central role in the translocation process. A series of ABCB1 isoforms containing a unique cysteine within TM6 was constructed and the resultant proteins purified and reconstituted. Accessibility of the cysteines to covalent modification by maleimide reagents was measured for the basal, ATP bound and vanadate trapped conformations of each isoform. Residues at the two extremes of the TM6 region examined (amino acids 344 to 360) were considerably more accessible than the central segment, the latter of which also failed to undergo significant conformational changes during the catalytic cycle. Covalent modification of the cytosolic segment of TM6 did, however, attenuate drug stimulation of ATP hydrolysis and demonstrates an important role for this segment in coupling drug binding to ATP hydrolysis during translocation.
dc.publisherAmerican Chemical Society
dc.sourceBiochemistry
dc.subjectKeywords: Cells; Drug delivery; Molecular mechanics; Oncology; Drug efflux pump; Intracellular drug accumulation; P-glycoprotein; Proteins; ABC transporter; adenosine triphosphate; coumarin; cysteine; glycoprotein P; maleimide; nicardipine; vanadic acid; vinblastin
dc.titleCytosolic region of TM6 in P-glycoprotein: Topographical analysis and functional perturbation by site directed labeling
dc.typeJournal article
local.description.notesImported from ARIES
local.identifier.citationvolume47
dc.date.issued2008
local.identifier.absfor060199 - Biochemistry and Cell Biology not elsewhere classified
local.identifier.absfor060110 - Receptors and Membrane Biology
local.identifier.absfor060112 - Structural Biology (incl. Macromolecular Modelling)
local.identifier.ariespublicationU3488905xPUB2574
local.identifier.ariespublicationu8801298xPUB81
local.type.statusPublished Version
local.contributor.affiliationStorm, Janet, University of Oxford
local.contributor.affiliationModok, Szabolcs, University of Oxford
local.contributor.affiliationO'Mara, Megan, University of Calgary
local.contributor.affiliationTieleman, D. Peter, University of Calgary
local.contributor.affiliationKerr, Ian, University of Nottingham
local.contributor.affiliationCallaghan, Richard, College of Medicine, Biology and Environment, ANU
local.description.embargo2037-12-31
local.bibliographicCitation.issue12
local.bibliographicCitation.startpage3615
local.bibliographicCitation.lastpage3624
local.identifier.doi10.1021/bi7023089
local.identifier.absseo970106 - Expanding Knowledge in the Biological Sciences
dc.date.updated2015-12-11T07:34:22Z
local.identifier.scopusID2-s2.0-41149140334
CollectionsANU Research Publications

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