Back to Search
Start Over
Modulation of constitutive activity and signaling bias of the ghrelin receptor by conformational constraint in the second extracellular loop.
- Source :
-
The Journal of biological chemistry [J Biol Chem] 2012 Sep 28; Vol. 287 (40), pp. 33488-502. Date of Electronic Publication: 2012 Jul 30. - Publication Year :
- 2012
-
Abstract
- Based on a rare, natural Glu for Ala-204(C+6) variant located six residues after the conserved Cys residue in extracellular loop 2b (ECL2b) associated with selective elimination of the high constitutive signaling of the ghrelin receptor, this loop was subjected to a detailed structure functional analysis. Introduction of Glu in different positions demonstrated that although the constitutive signaling was partly reduced when introduced in position 205(C+7) it was only totally eliminated in position 204(C+6). No charge-charge interaction partner could be identified for the Glu(C+6) variant despite mutational analysis of a number of potential partners in the extracellular loops and outer parts of the transmembrane segments. Systematic probing of position 204(C+6) with amino acid residues of different physicochemical properties indicated that a positively charged Lys surprisingly provided phenotypes similar to those of the negatively charged Glu residue. Computational chemistry analysis indicated that the propensity for the C-terminal segment of extracellular loop 2b to form an extended α-helix was increased from 15% in the wild type to 89 and 82% by introduction in position 204(C+6) of a Glu or a Lys residue, respectively. Moreover, the constitutive activity of the receptor was inhibited by Zn(2+) binding in an engineered metal ion site, stabilizing an α-helical conformation of this loop segment. It is concluded that the high constitutive activity of the ghrelin receptor is dependent upon flexibility in the C-terminal segment of extracellular loop 2 and that mutations or ligand binding that constrains this segment and thereby conceivably the movements of transmembrane domain V relative to transmembrane domain III inhibits the high constitutive signaling.
- Subjects :
- Alanine chemistry
Amino Acid Sequence
Animals
Arrestins metabolism
COS Cells
Chlorocebus aethiops
DNA Mutational Analysis
HEK293 Cells
Humans
Models, Biological
Models, Molecular
Molecular Sequence Data
Protein Conformation
Protein Structure, Secondary
Protein Structure, Tertiary
Receptors, G-Protein-Coupled chemistry
Receptors, Ghrelin chemistry
Signal Transduction
beta-Arrestins
Receptors, Ghrelin metabolism
Subjects
Details
- Language :
- English
- ISSN :
- 1083-351X
- Volume :
- 287
- Issue :
- 40
- Database :
- MEDLINE
- Journal :
- The Journal of biological chemistry
- Publication Type :
- Academic Journal
- Accession number :
- 22846991
- Full Text :
- https://doi.org/10.1074/jbc.M112.383240