| Title: | Double-mutant cycle scanning of the interaction of a peptide ligand and its G protein-coupled receptor |
| Author(s): | Naider F; Becker JM; Lee YH; Horovitz A; |
| Address: | "The College of Staten Island and Macromolecular Assemblies Institute of the City University of New York, Staten Island, New York 10314, USA. naider@mail.csi.cuny.edu" |
| ISSN/ISBN: | 0006-2960 (Print) 1520-4995 (Electronic) 0006-2960 (Linking) |
| Abstract: | "The interaction between the yeast G protein-coupled receptor (GPCR), Ste2p, and its alpha-factor tridecapeptide ligand was subjected to double-mutant cycle scanning analysis by which the pairwise interaction energy of each ligand residue with two receptor residues, N205 and Y266, was determined. The mutations N205A and Y266A were previously shown to result in deficient signaling but cause only a 2.5-fold and 6-fold decrease, respectively, in the affinity for alpha-factor. The analysis shows that residues at the amine terminus of alpha-factor interact strongly with N205 and Y266 whereas residues in the center and at the carboxyl terminus of the peptide interact only weakly if at all with these receptor residues. Multiple-mutant thermodynamic cycle analysis was used to assess whether the energies of selected pairwise interactions between residues of the alpha-factor peptide changed upon binding to Ste2p. Strong positive cooperativity between residues 1 through 4 of alpha-factor was observed during receptor binding. In contrast, no thermodynamic evidence was found for an interaction between a residue near the carboxyl terminus of alpha-factor (position 11) and one at the N-terminus (position 3). The study shows that multiple-mutant cycle analyses of the binding of an alanine-scanned peptide to wild-type and mutant GPCRs can provide detailed information on contributions of inter- and intramolecular interactions to the binding energy and potentially prove useful in developing 3D models of ligand docked to its receptor" |
| Keywords: | "Alanine/genetics/metabolism Amino Acid Sequence Binding, Competitive Ligands Mating Factor Peptides/*metabolism Receptors, Mating Factor/*genetics/*metabolism Saccharomyces cerevisiae Proteins/*genetics/*metabolism Thermodynamics Tyrosine/metabolism;" |
| Notes: | "MedlineNaider, Fred Becker, Jeffrey M Lee, Yong-Hun Horovitz, Amnon eng R01 GM022087/GM/NIGMS NIH HHS/ R01 GM022086-30/GM/NIGMS NIH HHS/ R01 GM022086/GM/NIGMS NIH HHS/ GM22086/GM/NIGMS NIH HHS/ GM22087/GM/NIGMS NIH HHS/ R56 GM022086/GM/NIGMS NIH HHS/ R01 GM022087-30/GM/NIGMS NIH HHS/ R56 GM022087/GM/NIGMS NIH HHS/ Research Support, N.I.H., Extramural 2007/02/15 Biochemistry. 2007 Mar 20; 46(11):3476-81. doi: 10.1021/bi602415u. Epub 2007 Feb 14" |
