title: Methods for recombinant expression and functional characterization of human cannabinoid receptor CB2.
abstract: Cannabinoid receptor CB2 is a seven transmembrane-domain integral membrane protein that belongs to a large superfamily of G protein-coupled receptors (GPCR). CB2 is a part of the endocannabinoid system that plays vital role in regulation of immune response, inflammation, pain sensitivity, obesity and other physiological responses. Information about the structure and mechanisms of functioning of this receptor in cell membranes is essential for the rational development of specific pharmaceuticals. Here we review the methodology for recombinant expression, purification, stabilization and biochemical characterization of CB2 suitable for preparation of multi-milligram quantities of functionally active receptor. The biotechnological protocols include expression of the recombinant CB2 in E. coli cells as a fusion with the maltose binding protein, stabilization with a high affinity ligand and a derivative of cholesterol in detergent micelles, efficient purification by tandem affinity chromatography, and reconstitution of the receptor into lipid bilayers. The purified recombinant CB2 receptor is amenable to functional and structural studies including nuclear magnetic resonance spectroscopy and a wide range of biochemical and biophysical techniques.
authors.affiliation: National Institute on Alcohol Abuse and Alcoholism, National Institutes of Health, 5625 Fishers Lane, Bethesda, MD 20892, USA.
authors.forename: Alexei A
authors.initials: AA
authors.lastname: Yeliseev
keywords: Cannabinoid receptor CB2; G protein activation; functional reconstitution; purification; recombinant expression