| Title: | Identification of residues of the Saccharomyces cerevisiae G protein-coupled receptor contributing to alpha-factor pheromone binding |
| Author(s): | Lee BK; Khare S; Naider F; Becker JM; |
| Address: | "Department of Microbiology, University of Tennessee, Knoxville, 37996, USA" |
| ISSN/ISBN: | 0021-9258 (Print) 0021-9258 (Linking) |
| Abstract: | "The Saccharomyces cerevisiae pheromone, alpha-factor (WHWLQLKPGQPMY), and Ste2p, its G protein-coupled receptor, were studied as a model for peptide ligand-receptor interaction. The affinities and activities of various synthetic position-10 alpha-factor analogs with Ste2p expressing mutations at residues Ser47 and Thr48 were investigated. All mutant receptors were expressed at a similar level in the cytoplasmic membrane, and their efficacies of signal transduction were similar to that of the wild-type receptor. Mutant receptors differed in binding affinity (Kd) and potency (EC50) for gene induction by alpha-factor. One mutant receptor (S47K,T48K) had dramatically reduced affinity and activity for [Lys10]- and [Orn10]alpha-factor, whereas the affinity for Saccharomyces kluyveri alpha-factor (WHWLSFSKGEPMY) was increased over 20-fold compared with that of wild-type receptor. In contrast, the affinity of [Lys10]- and [Orn10]alpha-factor was increased greatly in a S47E,T48E mutant receptor, whereas the binding of the S. kluyveri alpha-factor was abolished. The affinity of [Lys10]- and [Orn10]alpha-factor for the S47E,T48E receptor dropped 4-6-fold in the presence of 1 m NaCl, whereas the affinity of alpha-factor was not affected by this treatment. These results demonstrate that when bound to its receptor the 10th residue (Gln) of the S. cerevisiae alpha-factor is adjacent to Ser47 and Thr48 residues in the receptor and that the 10th residue of alpha-factors from two Saccharomyces species is responsible for the ligand selectivity to their cognate receptors. Based on these data, we have developed a two-dimensional model of alpha-factor binding to its receptor" |
| Keywords: | "Amino Acid Sequence Base Sequence Chromatography, High Pressure Liquid DNA Primers GTP-Binding Proteins/*metabolism Mating Factor Molecular Sequence Data Mutagenesis, Site-Directed Peptides/*metabolism Receptors, Cell Surface/chemistry/genetics/*metabolis;" |
| Notes: | "MedlineLee, B K Khare, S Naider, F Becker, J M eng GM22086/GM/NIGMS NIH HHS/ GM22087/GM/NIGMS NIH HHS/ Research Support, U.S. Gov't, P.H.S. 2001/08/10 J Biol Chem. 2001 Oct 12; 276(41):37950-61. doi: 10.1074/jbc.M103579200. Epub 2001 Aug 8" |
