Solubilization of serotonin1A receptors heterologously expressed in Chinese hamster ovary cells.
1. 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 behavioral and cognitive functions. 2. We report here, for the first time, the solubilization of 5-HT1A receptors stably expressed in Chinese Hamster Ovary (CHO) cells using the zwitterionic detergent CHAPS in presence of NaCl followed by polyethylene glycol (PEG) precipitation. We show by ligand-binding assay that the 5-HT1A receptor solubilized this way is functionally active. We have optimized the efficiency of solubilization with respect to total protein and NaCl concentration. 3. Our results show that careful control of salt and protein concentration is crucial in optimal solubilization of membrane receptors heterologously expressed in cells in culture. The effective solubilization of important neurotransmitter receptors such as 5-HT1A receptors which are present in very low amounts in the native tissue may represent an important step in characterizing membrane receptors expressed in mammalian cells in culture.
Chattopadhyay
Amitabha
A
Centre for Cellular and Molecular Biology, Hyderabad, India. amit@ccmb.res.in
Jafurulla
Md
M
Kalipatnapu
Shanti
S
D000818:Animals; D001667:Binding, Competitive / Q000502:physiology; D001681:Biological Assay; D016466:CHO Cells; D002462:Cell Membrane / Q000737:chemistry*; D011232:Chemical Precipitation; D002793:Cholic Acids / Q000737:chemistry; D006224:Cricetinae; D008954:Models, Biological; D009448:Neurochemistry / Q000379:methods*; D011092:Polyethylene Glycols / Q000737:chemistry; D044282:Receptor, Serotonin, 5-HT1A / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification*; D012701:Serotonin / Q000378:metabolism / Q000493:pharmacokinetics; D012965:Sodium Chloride / Q000737:chemistry / Q000494:pharmacology; D012995:Solubility / Q000187:drug effects; D009435:Synaptic Transmission / Q000502:physiology
D002793:Cholic Acids; D044282:Receptor, Serotonin, 5-HT1A; D012701:Serotonin; D011092:Polyethylene Glycols; D012965:Sodium Chloride; C028213:3-((3-cholamidopropyl)dimethylammonium)-1-propanesulfonate