Purification and characterization of the human ovarian LH/HCG receptor and comparison of the properties of mammalian LH/HCG receptors

Abstract

Methods previously published by us [Wimalasena et al., J Biol Chem 260: 10689–10697, 1985; Wimalasena et al., J Biol Chem 261: 9416–9420, 1986] were utilized to solubilize the human corpus luteal leuteinizing hormone/human chorionic gonadotropin (LH/hCG) receptor with 3-[(3-cholamide-propyl) dimethylammonio]-1-propanesulfonate (CHAPS) and to purify the receptor by two steps of hCG-Sepharose affinity chromatography. The specific binding capacity (SBC) of the purified human receptor was 7510 pmol/mg protein, and K a = 2.2 × 10 9 M −1 when iodo hCG was competed by hCG; the yield was 4–7% of starting activity. When hLH was used in competition with hCG, specific binding capacity was 7900 pmol/mg protein and K a 1.0 × 10 9 M −1 . Silver staining and autoradiography demonstrated a single protein of M r 78,000 under reducing and M r 58–62 × 10 3 under nonreducing conditions. Rat ovarian LH/hCG receptor was purified by similar methods and the K a of 3.5 × 10 10 M −1 for hCG was substantially different from the K a for hLH which was 2.1 × 10 9 M −1. M r of the rat protein was 78–82 × 10 3 (reduced) and 58–62 × 10 3 (nonreduced) when analyzed by silver staining and autoradiography. For the first time, human LH/hCG receptor has been purified to apparent homogen- eity, and its M r of 78,000 was essentially identical to the M r values of purified rat and porcine receptors.