Solubilization of high affinity G-protein-coupled serotonin1A receptors from bovine hippocampus using pre-micellar CHAPS at low concentration.
The serotonin1A (5-HT1A) receptors are members of a superfamily of seven transmembrane domain receptors that couple to G-proteins. They appear to be involved in various behavioural and cognitive functions. This paper reports an efficient strategy to solubilize 5-HT1A receptors from bovine hippocampal membranes using the zwitterionic detergent CHAPS which is mild and non-denaturing. Since high concentration of CHAPS has earlier been shown to induce dissociation and depletion of G-protein sub-units, a low (pre-micellar) concentration of CHAPS was used for solubilizing 5-HT1A receptors in the presence of NaCl followed by PEG precipitation. This results in solubilization of 5-HT1A receptors with a high degree of efficiency and gives rise to high affinity, functionally active G-protein-sensitive solubilized receptors. Optimal solubilization of the receptor from the native source with high ligand binding affinity and intact signal transduction components may constitute the first step in the molecular characterization of the 5-HT1A receptor in particular, and G-protein-coupled receptors in general.
Chattopadhyay
Amitabha
A
Centre for Cellular & Molecular Biology, Uppal Road, Hyderabad 500007, India. amit@ccmb.res.in
Harikumar
K G
KG
Kalipatnapu
Shanti
S
D017371:8-Hydroxy-2-(di-n-propylamino)tetralin / Q000378:metabolism; D000818:Animals; D001665:Binding Sites; D001667:Binding, Competitive; D002417:Cattle; D002462:Cell Membrane / Q000737:chemistry / Q000378:metabolism; D002499:Centrifugation, Density Gradient / Q000379:methods; D011232:Chemical Precipitation; D002793:Cholic Acids / Q000737:chemistry*; D003902:Detergents / Q000737:chemistry; D019204:GTP-Binding Proteins / Q000737:chemistry / Q000378:metabolism*; D006147:Guanine / Q000378:metabolism; D006624:Hippocampus / Q000737:chemistry*; D008823:Micelles; D008967:Molecular Biology / Q000379:methods*; D011092:Polyethylene Glycols / Q000737:chemistry; D011985:Receptors, Serotonin / Q000737:chemistry* / Q000378:metabolism*; D044263:Receptors, Serotonin, 5-HT1; D017366:Serotonin Receptor Agonists / Q000378:metabolism; D012965:Sodium Chloride / Q000737:chemistry; D012995:Solubility; D013395:Sucrose / Q000378:metabolism; D014316:Tritium
D002793:Cholic Acids; D003902:Detergents; D008823:Micelles; D011985:Receptors, Serotonin; D044263:Receptors, Serotonin, 5-HT1; D017366:Serotonin Receptor Agonists; D014316:Tritium; D011092:Polyethylene Glycols; D012965:Sodium Chloride; D013395:Sucrose; D006147:Guanine; D017371:8-Hydroxy-2-(di-n-propylamino)tetralin; D019204:GTP-Binding Proteins; C028213:3-((3-cholamidopropyl)dimethylammonium)-1-propanesulfonate