Identification of 70 calcium-sensing receptor mutations in hyper- and hypo-calcaemic patients: evidence for clustering of extracellular domain mutations at calcium-binding sites

Fadil M Hannan, M Andrew Nesbit, Chen Zhang, Treena Cranston, Alan J Curley, Brian Harding, Carl Fratter, Nigel Rust, Paul T Christie, Jeremy J O Turner, Manuel C Lemos, Michael R Bowl, Roger Bouillon, Caroline Brain, Nicola Bridges, Christine Burren, John M Connell, Heike Jung, Eileen Marks, David McCredieZulf Mughal, Christine Rodda, Sherida Tollefsen, Edward M Brown, Jenny J Yang, Rajesh V Thakker

Research output: Contribution to journalArticle (Academic Journal)peer-review

104 Citations (Scopus)

Abstract

The calcium-sensing receptor (CaSR) is a G-protein-coupled receptor that has an extracellular bilobed venus flytrap domain (VFTD) predicted to contain five calcium (Ca(2+))-binding sites. To elucidate the structure-function relationships of the VFTD, we investigated 294 unrelated probands with familial hypocalciuric hypercalcaemia (FHH), neonatal severe primary hyperparathyroidism (NSHPT) or autosomal dominant hypocalcaemic hypercalciuria (ADHH) for CaSR mutations and performed in vitro functional expression studies and three-dimensional modelling of mutations involving the VFTD. A total of 70 different CaSR mutations were identified: 35 in FHH, 10 in NSHPT and 25 in ADHH patients. Furthermore, a CaSR variant (Glu250Lys) was identified in FHH and ADHH probands and demonstrated to represent a functionally neutral polymorphism. NSHPT was associated with a large proportion of truncating CaSR mutations that occurred in the homozygous or compound heterozygous state. Thirty-four VFTD missense mutations were identified, and 18 mutations were located within 10 Å of one or more of the predicted Ca(2+)-binding sites, particularly at the VFTD cleft, which is the principal site of Ca(2+) binding. Mutations of residues 173 and 221, which are located at the entrance to the VFTD cleft binding site, were associated with both receptor activation (Leu173Phe and Pro221Leu) and inactivation (Leu173Pro and Pro221Gln), thereby highlighting the importance of these residues for entry and binding of Ca(2+) by the CaSR. Thus, these studies of disease-associated CaSR mutations have further elucidated the role of the VFTD cleft region in Ca(2+) binding and the function of the CaSR.

Original languageEnglish
Pages (from-to)2768-78
Number of pages11
JournalHuman Molecular Genetics
Volume21
Issue number12
DOIs
Publication statusPublished - 15 Jun 2012

Keywords

  • Binding Sites
  • Calcium
  • Genotype
  • HEK293 Cells
  • Humans
  • Hypercalcemia
  • Hyperparathyroidism
  • Hypocalcemia
  • Infant, Newborn
  • Models, Molecular
  • Mutation
  • Mutation Rate
  • Mutation, Missense
  • Protein Structure, Secondary
  • Protein Structure, Tertiary
  • Receptors, Calcium-Sensing
  • Journal Article
  • Research Support, Non-U.S. Gov't

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