Symmetric activation and modulation of the human calcium-sensing receptor

2021 ◽  
Vol 118 (51) ◽  
pp. e2115849118
Author(s):  
Jinseo Park ◽  
Hao Zuo ◽  
Aurel Frangaj ◽  
Ziao Fu ◽  
Laura Y. Yen ◽  
...  
Keyword(s):  
Transmembrane Domain ◽  
Allosteric Modulation ◽  
Allosteric Modulators ◽  
Calcium Sensing Receptor ◽  
Positive Allosteric Modulator ◽  
Functional Roles ◽  
Calcium Sensing ◽  
Positive Modulator ◽  
Key Residues ◽  
Subunit Contact

The human extracellular calcium-sensing (CaS) receptor controls plasma Ca2+ levels and contributes to nutrient-dependent maintenance and metabolism of diverse organs. Allosteric modulation of the CaS receptor corrects disorders of calcium homeostasis. Here, we report the cryogenic-electron microscopy reconstructions of a near–full-length CaS receptor in the absence and presence of allosteric modulators. Activation of the homodimeric CaS receptor requires a break in the transmembrane 6 (TM6) helix of each subunit, which facilitates the formation of a TM6-mediated homodimer interface and expansion of homodimer interactions. This transformation in TM6 occurs without a positive allosteric modulator. Two modulators with opposite functional roles bind to overlapping sites within the transmembrane domain through common interactions, acting to stabilize distinct rotamer conformations of key residues on the TM6 helix. The positive modulator reinforces TM6 distortion and maximizes subunit contact to enhance receptor activity, while the negative modulator strengthens an intact TM6 to dampen receptor function. In both active and inactive states, the receptor displays symmetrical transmembrane conformations that are consistent with its homodimeric assembly.

scholarly journals Negative allosteric modulators of the human calcium‐sensing receptor bind to overlapping and distinct sites within the 7‐transmembrane domain

2020 ◽  
Vol 177 (8) ◽  
pp. 1917-1930 ◽  
Author(s):  
Tracy M. Josephs ◽  
Andrew N. Keller ◽  
Elham Khajehali ◽  
Aaron DeBono ◽  
Christopher J. Langmead ◽  
...  
Keyword(s):  
Transmembrane Domain ◽  
Allosteric Modulators ◽  
Calcium Sensing Receptor ◽  
Calcium Sensing

scholarly journals Structural basis for activation and allosteric modulation of full-length calcium-sensing receptor

2021 ◽  
Vol 7 (23) ◽  
pp. eabg1483
Author(s):  
Tianlei Wen ◽  
Ziyu Wang ◽  
Xiaozhe Chen ◽  
Yue Ren ◽  
Xuhang Lu ◽  
...  
Keyword(s):  
Conformational Changes ◽  
Bound States ◽  
Hormone Secretion ◽  
Allosteric Modulation ◽  
Full Length ◽  
Structural Basis ◽  
Endocrine Disorders ◽  
Calcium Sensing Receptor ◽  
Calcium Sensing ◽  
Class C

Calcium-sensing receptor (CaSR) is a class C G protein–coupled receptor (GPCR) that plays an important role in calcium homeostasis and parathyroid hormone secretion. Here, we present multiple cryo–electron microscopy structures of full-length CaSR in distinct ligand-bound states. Ligands (Ca2+ and l-tryptophan) bind to the extracellular domain of CaSR and induce large-scale conformational changes, leading to the closure of two heptahelical transmembrane domains (7TMDs) for activation. The positive modulator (evocalcet) and the negative allosteric modulator (NPS-2143) occupy the similar binding pocket in 7TMD. The binding of NPS-2143 causes a considerable rearrangement of two 7TMDs, forming an inactivated TM6/TM6 interface. Moreover, a total of 305 disease-causing missense mutations of CaSR have been mapped to the structure in the active state, creating hotspot maps of five clinical endocrine disorders. Our results provide a structural framework for understanding the activation, allosteric modulation mechanism, and disease therapy for class C GPCRs.

scholarly journals Illuminating the allosteric modulation of the calcium-sensing receptor

2020 ◽  
Vol 117 (35) ◽  
pp. 21711-21722
Author(s):  
Hongkang Liu ◽  
Ping Yi ◽  
Wenjing Zhao ◽  
Yuling Wu ◽  
Francine Acher ◽  
...  
Keyword(s):  
Amino Acids ◽  
Conformational Changes ◽  
Resonance Energy Transfer ◽  
Resonance Energy ◽  
Membrane Receptors ◽  
Allosteric Modulators ◽  
Active Conformation ◽  
Calcium Sensing Receptor ◽  
Single Receptor ◽  
Calcium Sensing

Many membrane receptors are regulated by nutrients. However, how these nutrients control a single receptor remains unknown, even in the case of the well-studied calcium-sensing receptor CaSR, which is regulated by multiple factors, including ions and amino acids. Here, we developed an innovative cell-free Förster resonance energy transfer (FRET)-based conformational CaSR biosensor to clarify the main conformational changes associated with activation. By allowing a perfect control of ambient nutrients, this assay revealed that Ca2+alone fully stabilizes the active conformation, while amino acids behave as pure positive allosteric modulators. Based on the identification of Ca2+activation sites, we propose a molecular basis for how these different ligands cooperate to control CaSR activation. Our results provide important information on CaSR function and improve our understanding of the effects of genetic mutations responsible for human diseases. They also provide insights into how a receptor can integrate signals from various nutrients to better adapt to the cell response.

scholarly journals Widespread Changes in Positive Allosteric Modulation of the Muscarinic M1 Receptor in Some Participants With Schizophrenia

2019 ◽  
Vol 22 (10) ◽  
pp. 640-650 ◽  
Author(s):  
Shaun Hopper ◽  
Geoffrey Mark Pavey ◽  
Andrea Gogos ◽  
Brian Dean
Keyword(s):  
Carboxylic Acid ◽  
Radioligand Binding ◽  
Allosteric Modulation ◽  
Allosteric Modulator ◽  
Allosteric Modulators ◽  
Positive Allosteric Modulator ◽  
M1 Receptor ◽  
Area 6 ◽  
Subcortical Regions

Abstract Background Preclinical and some human data suggest allosteric modulation of the muscarinic M1 receptor (CHRM1) is a promising approach for the treatment of schizophrenia. However, it is suggested there is a subgroup of participants with schizophrenia who have profound loss of cortical CHRM1 (MRDS). This raises the possibility that some participants with schizophrenia may not respond optimally to CHRM1 allosteric modulation. Here we describe a novel methodology to measure positive allosteric modulation of CHRM1 in human CNS and the measurement of that response in the cortex, hippocampus, and striatum from participants with MRDS, non-MRDS and controls. Methods The cortex (Brodmann’s area 6), hippocampus, and striatum from 40 participants with schizophrenia (20 MRDS and 20 non-MRDS) and 20 controls were used to measure benzyl quinolone carboxylic acid-mediated shift in acetylcholine displacement of [3H]N-methylscopolamine using a novel in situ radioligand binding with autoradiography methodology. Results Compared with controls, participants with schizophrenia had lower levels of specific [3H]N-methylscopolamine binding in all CNS regions, whilst benzyl quinolone carboxylic acid-modulated binding was less in the striatum, Brodmann’s area 6, dentate gyrus, and subiculum. When divided by subgroup, only in MRDS was there lower specific [3H]N-methylscopolamine binding and less benzyl quinolone carboxylic acid-modulated binding in all cortical and subcortical regions studied. Conclusions In a subgroup of participants with schizophrenia, there is a widespread decreased responsiveness to a positive allosteric modulator at the CHRM1. This finding may have ramifications it positive allosteric modulators of the CHRM1 are used in clinical trials to treat schizophrenia as some participants may not have an optimal response.

scholarly journals Two novel mutations of the calcium-sensing receptor gene affecting the same amino acid position lead to opposite phenotypes and reveal the importance of p.N802 on receptor activity

2013 ◽  
Vol 168 (2) ◽  
pp. K27-K34 ◽  
Author(s):  
Anne-Sophie Lia-Baldini ◽  
Corinne Magdelaine ◽  
Angélique Nizou ◽  
Coraline Airault ◽  
Jean-Pierre Salles ◽  
...  
Keyword(s):  
Amino Acid ◽  
Transmembrane Domain ◽  
Direct Sequencing ◽  
Receptor Gene ◽  
Amino Acid Position ◽  
Receptor Activity ◽  
Site Directed Mutagenesis ◽  
Missense Mutations ◽  
Calcium Sensing Receptor ◽  
Calcium Sensing

ObjectiveGain-of-function mutations of the calcium-sensing receptor (CASR) gene have been identified in patients with sporadic or familial autosomal dominant hypocalcemia (ADH). Inactivating mutations of the CASR gene cause familial hypocalciuric hypercalcemia (FHH). Here, we report two novel CASR mutations affecting the same amino acid (p.N802); one causes ADH and the other atypical FHH.Patients and methodsThe first patient, an 11-year-old girl suffering from hypocalcemia, developed nephrocalcinosis when she was only 5 years old. The second patient is a 30-year-old woman who presented with mild hypercalcemia. PCR amplification of CASR coding exons and direct sequencing of PCR products were used to identify mutations. Site-directed mutagenesis was used to generate mutated CASR cDNAs in an expression plasmid. Using the MAPK assay system and transient transfection of Cos-7 cells with wild-type (WT) and mutated CASR, we studied the responses of these mutated receptors to extracellular Ca2+ and to the negative allosteric CASR modulator, NPS2143.ResultsTwo heterozygous missense mutations (p.N802I and p.N802S) affecting a residue in the sixth transmembrane domain of CASR were identified. In functional tests, the response of the p.N802S mutant to calcium was typical of an inactivating mutation. However, the p.N802I mutant had 70% of the maximally stimulated WT receptor activity even in the absence of extracellular calcium. This constitutive activity was only partially inhibited by the inhibitor, NPS2143.ConclusionsThe asparagine at amino acid position 802 appears to be essential for the activity of the CASR protein and is implicated in the mechanism of CASR signaling.

scholarly journals Divergent effects of strontium and calcium-sensing receptor positive allosteric modulators (calcimimetics) on human osteoclast activity

2018 ◽  
Vol 175 (21) ◽  
pp. 4095-4108 ◽  
Author(s):  
Natalie A Diepenhorst ◽  
Katie Leach ◽  
Andrew N Keller ◽  
Patricia Rueda ◽  
Anna E Cook ◽  
...  
Keyword(s):  
Allosteric Modulators ◽  
Calcium Sensing Receptor ◽  
Osteoclast Activity ◽  
Calcium Sensing ◽  
Human Osteoclast ◽  
Positive Allosteric Modulators

scholarly journals Homology Modeling of the Transmembrane Domain of the Human Calcium Sensing Receptor and Localization of an Allosteric Binding Site

2003 ◽  
Vol 279 (8) ◽  
pp. 7254-7263 ◽  
Author(s):  
Susanne U. Miedlich ◽  
Lucio Gama ◽  
Klaus Seuwen ◽  
Romain M. Wolf ◽  
Gerda E. Breitwieser
Keyword(s):  
Homology Modeling ◽  
Binding Site ◽  
Transmembrane Domain ◽  
Calcium Sensing Receptor ◽  
Calcium Sensing

scholarly journals Loss-of-Function and Gain-of-Function Mutations of Calcium-Sensing Receptor: Functional Analysis and the Effect of Allosteric Modulators NPS R-568 and NPS 2143

2013 ◽  
Vol 98 (10) ◽  
pp. E1692-E1701 ◽  
Author(s):  
Akie Nakamura ◽  
Tomoyuki Hotsubo ◽  
Keiji Kobayashi ◽  
Hiroshi Mochizuki ◽  
Katsura Ishizu ◽  
...  
Keyword(s):  
Cell Surface ◽  
Surface Expression ◽  
Cell Surface Expression ◽  
Allosteric Modulators ◽  
Familial Hypocalciuric Hypercalcemia ◽  
Calcium Sensing Receptor ◽  
Activating Mutations ◽  
Casr Gene ◽  
Calcium Sensing ◽  
Inactivating Mutations

Abstract Objective: Activating mutations in the calcium-sensing receptor (CASR) gene cause autosomal dominant hypoparathyroidism, and heterozygous inactivating CASR mutations cause familial hypocalciuric hypercalcemia. Recently, there has been a focus on the use of allosteric modulators to restore the functional activity of mutant CASRs. In this study, the effect of allosteric modulators NPS R-568 and NPS 2143 on CASR mutants was studied in vitro. Methods: DNA sequence analysis of the CASR gene was undertaken in autosomal dominant hypoparathyroidism and familial hypocalciuric hypercalcemia Japanese patients, and the functional consequences for the Gi-MAPK pathway and cell surface expression of CASR were determined. Furthermore, we studied the effect of NPS R-568 and NPS 2143 on the signal transduction activity and cell surface expression of each mutant CASR. Results: We identified 3 activating mutations (S122C, P569H, and I839T) and 2 inactivating mutations (A110T and R172G) in patients. The activating and inactivating mutations caused leftward and rightward shifts, respectively, in the dose-response curves of the signaling pathway. NPS R-568 rescued the signal transduction capacity of 2 inactivating mutants without increasing cell surface expression levels. NPS 2143 suppressed the enhanced activity of the activating mutants without altering cell surface expression levels, although A843E, which is a constitutively active mutant, was suppressed to a lesser degree. Conclusions: We have identified 4 novel mutations of CASR. Moreover, our results indicate that allosteric modulators can restore the activity of the loss- and gain-of-function mutant CASRs, identified in this study.

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