Display Settings:

Format

Send to:

Choose Destination
See comment in PubMed Commons below
J Mol Biol. 2011 Jun 17;409(4):513-28. doi: 10.1016/j.jmb.2011.03.059. Epub 2011 Apr 6.

Differential interactions of fluorescent agonists and antagonists with the yeast G protein coupled receptor Ste2p.

Author information

  • 1Department of Biochemistry and Biophysics, University of Rochester School of Medicine and Dentistry, Rochester, NY 14642, USA.

Abstract

We describe a rapid method to probe for mutations in cell surface ligand-binding proteins that affect the environment of bound ligand. The method uses fluorescence-activated cell sorting to screen randomly mutated receptors for substitutions that alter the fluorescence emission spectrum of environmentally sensitive fluorescent ligands. When applied to the yeast α-factor receptor Ste2p, a G protein-coupled receptor, the procedure identified 22 substitutions that red shift the emission of a fluorescent agonist, including substitutions at residues previously implicated in ligand binding and at additional sites. A separate set of substitutions, identified in a screen for mutations that alter the emission of a fluorescent α-factor antagonist, occurs at sites that are unlikely to contact the ligand directly. Instead, these mutations alter receptor conformation to increase ligand-binding affinity and provide signaling in response to antagonists of normal receptors. These results suggest that receptor--agonist interactions involve at least two sites, of which only one is specific for the activated conformation of the receptor.

Copyright © 2011. Published by Elsevier Ltd.

PMID:
21477594
[PubMed - indexed for MEDLINE]
PMCID:
PMC3104124
Free PMC Article
PubMed Commons home

PubMed Commons

0 comments
How to join PubMed Commons

    Supplemental Content

    Icon for Elsevier Science Icon for PubMed Central
    Loading ...
    Write to the Help Desk