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Biochemistry. 2014 Jan 14;53(1):4-6. doi: 10.1021/bi401412e. Epub 2013 Dec 30.

Radioligand binding to nanodisc-reconstituted membrane transporters assessed by the scintillation proximity assay.

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1
Department of Cellular and Molecular Physiology, Yale University School of Medicine , New Haven, Connecticut 06520, United States.

Erratum in

  • Biochemistry. 2014 Feb 25;53(7):1239.

Abstract

The scintillation proximity assay is a powerful technique for measuring radioligand binding to membrane transporters and has become an integral part of high-throughput drug discovery screening efforts. Here we adapt the method for use with purified LeuT, a prokaryotic secondary transporter, reconstituted into phospholipid bilayer nanodiscs. This application surmounts potential challenges with background interference from endogenously expressed proteins, aggregation and loss of binding activity often accompanying detergent solubilization from native cell membranes, and heterogeneity in size and transporter orientation, where at least some ligand binding sites are inaccessible, associated with reconstitution into lipid vesicles.

PMID:
24344975
PMCID:
PMC4062192
DOI:
10.1021/bi401412e
[Indexed for MEDLINE]
Free PMC Article
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