A serotonin 5-HT3 receptor was functionally expressed to high levels and on a large scale in mammalian cells with the Semliki Forest virus system. Conditions were optimized to maximize detergent solubilization of the receptor, while preserving ligand binding activity. An efficient one-step purifn. yielding .apprx.50% of the histidine-tagged 5-HT3 receptor was achieved with immobilized metal ion chromatog. The expressed receptor, in both membranes and purified prepns., exhibited wild-type ligand binding properties, characterized by one class of binding sites. The purity of the receptor was shown by SDS-PAGE, yielding a single band at 65 kDa, and was confirmed by the specific ligand binding activity of .apprx.5 nmol/mg of protein. Deglycosylation of the receptor reduced the estd. relative mol. mass to 49 kDa. The apparent mol. mass of the functional receptor complex was detd. by size exclusion chromatog. to be 280 kDa, suggesting that the 5-HT3 receptor is a pentameric homooligomer. The secondary structure of the 5-HT3 receptor as detd. by CD appeared to consist of mainly a-helixes (50%) and b-strands (24%), with minor contributions from nonregular structure (9%). The binding of either agonist or antagonist did not alter the secondary structure of the receptor. [on SciFinder (R)]