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Role of protonmotive-force and ATP-binding/hydrolysis in multidrug translocation by the ATP-binding cassette transporter LmrA

Reference	BB/C004663/1
Principal Investigator / Supervisor	Professor Hendrik Willem van Veen
Co-Investigators / Co-Supervisors
Institution	University of Cambridge
Department	Pharmacology
Funding type	Research
Value (£)	199,279
Status	Completed
Type	Research Grant
Start date	14/04/2005
End date	13/09/2008
Duration	41 months

Abstract

Multidrug transporters mediate the extrusion of structurally unrelated drugs from prokaryotic and eukaryotic cells. As a result of this efflux activity, the cytoplasmic drug concentration in the cell is lowered to subtoxic levels, and hence, cells become multidrug resistant. Multidrug transporters interfere with the chemotherapeutic treatment of cancers and infections by pathogenic microorganisms. This research proposal focuses on the multidrug transporter LmrA, a bacterial homologue of the human multidrug resistance P-glycoprotein that can functionally replace its human counterpart in human lung fibroblast cells [Nature 391: 291-295 (1998)]. LmrA is an ATP-binding cassette (ABC) transporter containing one membrane domain with 6 transmembrane spanning segments and one nucleotide-binding domain. LmrA homodimerises to form a functional transporter with the 4 core domains [EMBO J. 19: 2503-2514 (2000)]. Surprisingly, our recent observations suggest that the membrane domain of LmrA (LmrA-MD) is functional as a reversible, protonmotive force-driven secondary-active multidrug uptake system, which, in full-length LmrA, acts in conjunction with the nucleotide-binding domain to enable ATP-dependent efflux of drugs and protons in bacterial cells [Nature 426: 866-869 (2003)]. We also identified an acidic glutamate (glutamate-314) in the intracellular domain of LmrA that appears to be required for LmrA-mediated proton translocation [Nature 426: 866-869 (2003)]. The reversibility of drug transport by LmrA-MD is maintained in LmrA [J. Biol. Chem. 279: 11273-11280 (2004)]. This project will focus in more detail on the role of the protonmotive-force and ATP-binding/hydrolysis in LmrA-mediated drug translocation. The project entails a multidisciplinary strategy involving molecular biology-based methods as well as biochemical and biophysical techniques. This strategy will add important new information about the molecular mechanism of this unique class of MDR transporters, and may lead to the development of new modulators that could induce drug uptake by ABC-type multidrug transporters in drug-resistant cells through their ability to uncouple ATP binding/hydrolysis from multidrug efflux.

Summary

unavailable

Committee	Closed Committee - Biochemistry & Cell Biology (BCB)
Research Topics	Microbiology, Pharmaceuticals, Structural Biology
Research Priority	X – Research Priority information not available
Research Initiative	X - not in an Initiative
Funding Scheme	X – not Funded via a specific Funding Scheme

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