title: Solubilization of luteinizing hormone receptor from human corpora lutea in a stable form and identification of the hormone-binding unit by ligand blotting.
abstract: LH receptors were solubilized from human corpora lutea in phosphate-buffered saline containing 1% Triton X-100, 20% glycerol, and protease inhibitors. The presence of 20% (vol/vol) glycerol was necessary for quantitative preservation of [125I]hCG-binding activity in detergent solution. The solubilized receptors were stable for several weeks at -20 C and at -80 C and for at least 18 h at 4 C. Binding of [125I]hCG to the soluble LH receptors was time and temperature dependent and varied linearly with the amount of soluble protein. Equilibrium binding studies revealed a single class of high affinity [125I]hCG-binding sites with an equilibrium dissociation constant (Kd) of 4.3 x 10(-10) mol/L (at 20 C). The molecular size of the human LH receptors was analyzed by ligand blotting. Solubilized receptors were subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis under nonreducing conditions and transferred to nitrocellulose. Incubation of the protein blot with [125I]hCG to the 85/90K mol wt species was inhibited by unlabeled hCG. These results indicate that LH receptors can be solubilized in nonionic detergent while maintaining their hormone-binding activity and demonstrate that the receptors contain 85/90K hormone-binding species.
authors.affiliation: Biocenter, University of Oulu, Finland.
authors.forename: K P
authors.initials: KP
authors.lastname: Keinänen
authors.forename: H J
authors.initials: HJ
authors.lastname: Rajaniemi
mesh_terms: D001665:Binding Sites / Q000187:drug effects; D003338:Corpus Luteum / Q000032:analysis*; D003902:Detergents / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D005260:Female; D005990:Glycerol / Q000494:pharmacology; D006801:Humans; D008024:Ligands; D011974:Receptors, LH / Q000302:isolation & purification*; D012995:Solubility
chemical_list: D003902:Detergents; D008024:Ligands; D011974:Receptors, LH; D005990:Glycerol