Highly Conserved Serine in the Third Transmembrane Helix of the Luteinizing Hormone/Human Chorionic Gonadotropin Receptor Regulates Receptor Activation

Abstract
The elucidation of the role of highly conserved polar amino acids in the transmembrane helices of G-protein-coupled receptors (GPCRs) is important in understanding the mechanism of receptor activation. To this end, the significance of a highly conserved serine residue in the third transmembrane α-helix (TM3) of the luteinizing hormone/human chorionic gonadotropin receptor (LH/hCGR) in regulating receptor activation was examined. Results showed that mutation of serine 431 to alanine (S431A) decreased the ability of the receptor to mediate cAMP production in response to hCG, suggesting that S431 stabilizes the active state of the receptor. Homology with other GPCRs suggests that S431 may participate in the coordination of a Na+ ion. Since Na+ has been found to stabilize the active state of the receptor in the presence of hCG, the possibility that S431 promotes receptor activation by mediating the effects of Na+ was explored. Results showed that the regulation of hormone-induced receptor activation by S431 was independent of Na+. A rhodopsin-based homology model of the TM region of the LH/hCGR was developed to identify other amino acids that might mediate the effects of Na+ on receptor function. Results indicate that substitution of an Asp at position 556 with Tyr alters the ability of Na+ to regulate receptor activation. The homology model is used to explain this result as well as to identify a mechanism through which S431 may regulate receptor signaling. Taken together, these studies provide novel insights into the mechanism of LH/hCG receptor activation.