Delineating a Ca2+ binding pocket within the venus flytrap module of the human calcium-sensing receptor.
The Ca(2+)-sensing receptor (CaSR) belongs to the class III G-protein-coupled receptors (GPCRs), which include receptors for pheromones, amino acids, sweeteners, and the neurotransmitters glutamate and gamma-aminobutyric acid (GABA). These receptors are characterized by a long extracellular amino-terminal domain called a Venus flytrap module (VFTM) containing the ligand binding pocket. To elucidate the molecular determinants implicated in Ca(2+) recognition by the CaSR VFTM, we developed a homology model of the human CaSR VFTM from the x-ray structure of the metabotropic glutamate receptor type 1 (mGluR1), and a phylogenetic analysis of 14 class III GPCR VFTMs. We identified critical amino acids delineating a Ca(2+) binding pocket predicted to be adjacent to, but distinct from, a cavity reminiscent of the binding site described for amino acids in mGluRs, GABA-B receptor, and GPRC6a. Most interestingly, these Ca(2+)-contacting residues are well conserved within class III GPCR VFTMs. Our model was validated by mutational and functional analysis, including the characterization of activating and inactivating mutations affecting a single amino acid, Glu-297, located within the proposed Ca(2+) binding pocket of the CaSR and associated with autosomal dominant hypocalcemia and familial hypocalciuric hypercalcemia, respectively, genetic diseases characterized by perturbations in Ca(2+) homeostasis. Altogether, these data define a Ca(2+) binding pocket within the CaSR VFTM that may be conserved in several other class III GPCRs, thereby providing a molecular basis for extracellular Ca(2+) sensing by these receptors.
Silve
Caroline
C
INSERM, U426, Faculté deMédecine Xavier Bichat and IFR02, Paris, France.
Petrel
Christophe
C
Leroy
Christine
C
Bruel
Henri
H
Mallet
Eric
E
Rognan
Didier
D
Ruat
Martial
M
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D002118:Calcium / Q000378:metabolism*; D005260:Female; D006801:Humans; D008297:Male; D009154:Mutation; D010375:Pedigree; D010802:Phylogeny; D051381:Rats; D018091:Receptors, AMPA / Q000378:metabolism; D044169:Receptors, Calcium-Sensing / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*
D002118:Calcium; D018091:Receptors, AMPA; D044169:Receptors, Calcium-Sensing