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

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

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.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Transport Systems, Neutral / analysis*
  • Amino Acid Transport Systems, Neutral / metabolism*
  • Bacterial Proteins / analysis*
  • Bacterial Proteins / metabolism*
  • Ligands
  • Lipid Bilayers / metabolism
  • Nanostructures / chemistry
  • Scintillation Counting*

Substances

  • Amino Acid Transport Systems, Neutral
  • Bacterial Proteins
  • Ligands
  • Lipid Bilayers