Roles of transmembrane prolines and proline-induced kinks of the lutropin/choriogonadotropin receptor

The lutropin/choriogonadotropin receptor is a seven-helix transmembrane (TM) receptor, A unique feature of TM helices is the content of Pro, which generally is absent in alpha helices of globular proteins, Because Pro disrupts helices and introduces a similar to 26 degrees kink, it has been speculated that Pro plays a crucial role in the structure of TM helices, exoloops, and cytoloops of TM receptors. To examine the roles of the five TM Pros of the lutropin/choriogonadotropin receptor, these residues were individually substituted, Mutant receptors were examined for surface expression, hormone binding, and cAMP induction, Surface expression was monitored after introducing the flag epitope into the receptors. Flag epitopes slightly affected cAMP induction but not hormone binding or surface expression of receptors as monitored by immunofluorescence microscopy and I-125-anti-flag antibody, The results indicate that Pro(479) in TM 4 and pro(598) in TM 7 play important yet contrasting roles. pro(479) is crucial for hormone binding at the cell surface but not after solubilization of the receptor. This is more likely due to the Pro side chain than the Pro-induced kink. pro(598) is important for surface expression. The kinks of pro(463) of TM 4, Pro(562) of TM 6, or pro(591) of TM 7 are not important because the substitution of Phe for these residues did not significantly impact surface expression, hormone binding, and cAMP induction.