Changes in conformation at the cytoplasmic ends of the fifth and sixth transmembrane helices of a yeast G protein-coupled receptor in response to ligand binding.

摘要: The third intracellular loop (IL3) of G protein-coupled receptors (GPCRs) is an important contact domain between GPCRs and their proteins. Previously, the IL3 Ste2p, a Saccharomyces cerevisiae GPCR, was suggested to undergo conformational change upon activation as detected by differential protease susceptibility in presence absence ligand. In this study using disulfide cross-linking experiments we show that Ste2p cytoplasmic ends helix 5 (TM5) 6 (TM6) flank amino carboxyl sides changes ligand binding, whereas center does not. Single Cys substitution residues middle led formed high levels cross-linked at interface contiguous TM5 TM6 resulted minimal disulfide-mediated receptor. alternating pattern involved 310 IL3. Agonist (WHWLQLKPGQPNleY) induced reduced mediated substitutions but not Thus, binding. An α-factor antagonist (des-Trp, des- His-α-factor) did influence cross-linking, suggesting interaction N-terminus α- factor with critical for inducing TM6. We propose conformation revealed are affected protein activation. This provides new information about role specific GPCR signal transduction how peptide binding activates