This Article Full Text (PDF) Submit a related Letter to the Editor Alert me when this article is cited Alert me when eLetters are posted Alert me if a correction is posted Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Request Copyright Permission Citing Articles Citing Articles via Google Scholar Google Scholar Articles by KEINÄNEN, K. P. Articles by RAJANIEMI, H. J. Search for Related Content PubMed PubMed Citation Articles by KEINÄNEN, K. P. Articles by RAJANIEMI, H. J.

Solubilization of Luteinizing Hormone Receptor From Human Corpora Lutea in a Stable Form and Identification of the Hormone-Binding Unit by Ligand Blotting^*

Biocenter and Department of Anatomy, University of Oulu Oulu, Finland

Address all correspondence and requests for reprints to: Kari P. Keinanen, Department of Anatomy, University of Oulu, Kajaanintie 52 A, SF-90220 Oulu, Finland.

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 (K_d) of 4.3 x l0^–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 followed by autoradiography revealed a broad 90K mol wt band or, in some samples, an 85/90K mol wt doublet. Binding of [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.

^* This work was supported by the Academy of Finland, the Cultural Foundation of Finland, and Emil Aaltonen Foundation.

Received October 29, 1987.