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  • Conformational antibody binding to a native, cell-free expressed GPCR in block copolymer membranes.

Conformational antibody binding to a native, cell-free expressed GPCR in block copolymer membranes.

PloS one (2014-10-21)
Hans-Peter M de Hoog, Esther M Lin JieRong, Sourabh Banerjee, Fabien M Décaillot, Madhavan Nallani
ABSTRACT

G-protein coupled receptors (GPCRs) play a key role in physiological processes and are attractive drug targets. Their biophysical characterization is, however, highly challenging because of their innate instability outside a stabilizing membrane and the difficulty of finding a suitable expression system. We here show the cell-free expression of a GPCR, CXCR4, and its direct embedding in diblock copolymer membranes. The polymer-stabilized CXCR4 is readily immobilized onto biosensor chips for label-free binding analysis. Kinetic characterization using a conformationally sensitive antibody shows the receptor to exist in the correctly folded conformation, showing binding behaviour that is commensurate with heterologously expressed CXCR4.

MATERIALS
Product Number
Brand
Product Description

Aucubin, primary reference standard
Sigma-Aldrich
Ethanolamine, liquid, BioReagent, suitable for cell culture, ≥98%
Sigma-Aldrich
Ethanolamine, ≥98%
Sigma-Aldrich
Ethanolamine, ≥99%
Supelco
Aucubin, analytical standard
Supelco
Ethanolamine, analytical standard
Sigma-Aldrich
Ethanolamine, purified by redistillation, ≥99.5%
Sigma-Aldrich
Ethanolamine, ACS reagent, ≥99.0%
Sigma-Aldrich
Ethanolamine, puriss. p.a., ACS reagent, ≥99.0% (GC/NT)
Trolamine impurity A, European Pharmacopoeia (EP) Reference Standard