scholarly journals Ligand recognition and G-protein coupling selectivity of cholecystokinin A receptor

2021 ◽  
Author(s):  
Qiufeng Liu ◽  
Dehua Yang ◽  
Youwen Zhuang ◽  
Tristan I. Croll ◽  
Xiaoqing Cai ◽  
...  
Keyword(s):  
G Protein ◽  
Metabolic Diseases ◽  
Ligand Recognition ◽  
G Protein Coupling ◽  
Protein Coupling ◽  
Cryo Electron Microscopy ◽  
Receptor Recognition ◽  
G Protein Coupled ◽  
Nutrient Homeostasis ◽  
Similar Conformation

AbstractCholecystokinin A receptor (CCKAR) belongs to family A G-protein-coupled receptors and regulates nutrient homeostasis upon stimulation by cholecystokinin (CCK). It is an attractive drug target for gastrointestinal and metabolic diseases. One distinguishing feature of CCKAR is its ability to interact with a sulfated ligand and to couple with divergent G-protein subtypes, including Gs, Gi and Gq. However, the basis for G-protein coupling promiscuity and ligand recognition by CCKAR remains unknown. Here, we present three cryo-electron microscopy structures of sulfated CCK-8-activated CCKAR in complex with Gs, Gi and Gq heterotrimers, respectively. CCKAR presents a similar conformation in the three structures, whereas conformational differences in the ‘wavy hook’ of the Gα subunits and ICL3 of the receptor serve as determinants in G-protein coupling selectivity. Our findings provide a framework for understanding G-protein coupling promiscuity by CCKAR and uncover the mechanism of receptor recognition by sulfated CCK-8.

scholarly journals Structural basis for ligand recognition and G protein-coupling promiscuity of the cholecystokinin A receptor

2021 ◽  
Author(s):  
Qiufeng Liu ◽  
Dehua Yang ◽  
Youwen Zhuang ◽  
Tristan I Croll ◽  
Xiaoqing Cai ◽  
...  
Keyword(s):  
G Protein ◽  
Metabolic Diseases ◽  
Structural Basis ◽  
Ligand Recognition ◽  
G Protein Coupling ◽  
Protein Coupling ◽  
Cryo Electron Microscopy ◽  
G Protein Coupled ◽  
Nutrient Homeostasis ◽  
Similar Conformation

Cholecystokinin A receptor (CCKAR) belongs to family A G protein-coupled receptors (GPCRs) and regulates nutrient homeostasis upon stimulation by cholecystokinin (CCK). It is an attractive drug target for gastrointestinal and metabolic diseases. One distinguishing feature of CCKAR is its ability to interact with sulfated ligand and to couple with divergent G protein subtypes, including Gs, Gi, and Gq. However, the basis for G protein coupling promiscuity and ligand recognition by CCKAR remain unknown. Here we present three cryo-electron microscopy (cryo-EM) structures of sulfated CCK-8 activated CCKAR in complex with Gs, Gi, and Gq heterotrimers, respectively. In these three structures, CCKAR presents a similar conformation, whereas conformational differences in ″wavy hook″ of Gα subunits and ICL3 of the receptor serve as determinants in G protein coupling selectivity. These structures together with mutagenesis data provide the framework for understanding the G protein coupling promiscuity by CCKAR and uncover the mechanism of receptor recognition by sulfated CCK-8.

Oxidative stress reduces renal dopamine D1 receptor-Gq/11α G protein-phospholipase C signaling involving G protein-coupled receptor kinase 2

2007 ◽  
Vol 293 (1) ◽  
pp. F306-F315 ◽  
Author(s):  
Anees Ahmad Banday ◽  
Mustafa F. Lokhandwala
Keyword(s):  
Oxidative Stress ◽  
Atpase Activity ◽  
Phospholipase C ◽  
G Protein ◽  
Lipoic Acid ◽  
Skf 38393 ◽  
G Protein Coupling ◽  
Protein Coupling ◽  
Obese Rats ◽  
G Protein Coupled

The dopamine D1 receptors (D1R), expressed in renal proximal tubules, participate in the regulation of sodium transport. A defect in the coupling of the D1R to its G protein/effector complex in renal tubules has been reported in various conditions associated with oxidative stress. Because G protein-coupled receptor kinases (GRKs) are known to play an important role in D1R desensitization, we tested the hypothesis that increased oxidative stress in obese Zucker rats may cause GRK2 upregulation and, subsequently, D1R dysfunction. Lean and obese rats were given normal diet or diet supplemented with antioxidant lipoic acid for 2 wk. Compared with lean rats, obese rats exhibited oxidative stress, D1R were uncoupled from Gq/11α at basal level, and SKF-38393 failed to elicit D1R-G protein coupling, stimulate phospholipase C (PLC), and inhibit Na-K-ATPase activity. These animals showed increased basal protein kinase C (PKC) activity and membranous translocation of GRK2 and increased GKR2-Gq/11α interaction and D1R serine phosphorylation. Enzymatic dephosphorylation of D1R restored SKF-38393-induced adenylyl cyclase stimulation but not PLC activation. Treatment of obese rats with lipoic acid restored D1R-G protein coupling and SKF-38393-induced PLC stimulation and Na-K-ATPase inhibition. Lipoic acid treatment also normalized PKC activity, GRK2 sequestration, and GKR2-Gq/11α interaction. In conclusion, these data show that oxidative stress increases PKC activity causing GRK2 membranous translocation. GRK2 interacts with Gq/11α and acts, at least in part, as a regulator of G protein signaling leading to the D1R-Gq/11α uncoupling, causing inability of SKF-38393 to stimulate PLC and inhibit Na/K-ATPase. Lipoic acid, while reducing oxidative stress, normalized PKC activity and restored D1R-Gq/11α-PLC signaling and the ability of SKF-38393 to inhibit Na-K-ATPase activity.

Focus Issue: New insights in GPCR to G protein signaling

2016 ◽  
Vol 9 (423) ◽  
pp. eg6-eg6 ◽  
Author(s):  
Nancy R. Gough
Keyword(s):  
G Protein ◽  
G Protein Signaling ◽  
Constitutive Activity ◽  
Protein Signaling ◽  
Polarized Growth ◽  
G Protein Coupling ◽  
Α Subunit ◽  
Protein Coupling ◽  
G Protein Coupled ◽  
Orphan Gpcr

This Focus Issue highlights new discoveries at the level of the receptor, the α subunit, and the βγ subunit and spans research in yeast on polarized growth and G protein–coupled receptor (GPCR) trafficking, in mice on an orphan GPCR with constitutive activity, and a disease-causing mutation in an α subunit that results in inappropriate GPCR–G protein coupling.

scholarly journals Using Ligand-Induced Conformational Change to Screen for Compounds Targeting G-Protein-Coupled Receptors

2007 ◽  
Vol 12 (2) ◽  
pp. 175-185 ◽  
Author(s):  
Brian F. O'Dowd ◽  
Mohammad Alijaniaram ◽  
Xiaodong Ji ◽  
Tuan Nguyen ◽  
Richard M. Eglen ◽  
...  
Keyword(s):  
Cell Surface ◽  
G Protein ◽  
High Throughput Screening ◽  
G Protein Coupled Receptors ◽  
Nuclear Localization Sequence ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
G Protein Coupling ◽  
Protein Coupling ◽  
G Protein Coupled

The authors describe a novel drug strategy designed as a primary screen to discover either antagonist or agonist compounds targeting G-protein-coupled receptors (GPCRs). The incorporation of a nuclear localization sequence (NLS, a 5 amino acid substitution), in a location in helix 8 of the GPCR structure, resulted in ligand-independent receptor translocation from the cell surface to the nucleus. Blockade of the GPCR-NLS translocation from the cell surface was achieved by either antagonist or agonist treatments, each achieving their result in a sensitive concentration-dependent manner. GPCR-NLS translocation and blockade occurred regardless of the identity of the G-protein-coupling, and thus this assay is also ideally suited for identification of compounds targeting orphan GPCRs. The GPCR-NLS trafficking was visualized by fusion to fluorescent detectable proteins. Quantification of this effect was measured by determining the density of cell surface receptors, using enzyme fragment complementation in a manner suitable for high-throughput screening. Thus, the authors have developed a cellular assay for GPCRs suitable for compound screening without requiring prior identification of an agonist or knowledge of G-protein-coupling.

scholarly journals Analysis of β2AR-Gs and β2AR-Gi complex formation by NMR spectroscopy

2020 ◽  
Vol 117 (37) ◽  
pp. 23096-23105 ◽  
Author(s):  
Xiuyan Ma ◽  
Yunfei Hu ◽  
Hossein Batebi ◽  
Jie Heng ◽  
Jun Xu ◽  
...  
Keyword(s):  
Nmr Spectroscopy ◽  
Complex Formation ◽  
G Protein ◽  
Conformational Changes ◽  
Structural Changes ◽  
Protein G ◽  
G Protein Coupling ◽  
Protein Coupling ◽  
G Protein Coupled ◽  
Camp Formation

The β2-adrenergic receptor (β2AR) is a prototypical G protein-coupled receptor (GPCR) that preferentially couples to the stimulatory G protein Gs and stimulates cAMP formation. Functional studies have shown that the β2AR also couples to inhibitory G protein Gi, activation of which inhibits cAMP formation [R. P. Xiao, Sci. STKE 2001, re15 (2001)]. A crystal structure of the β2AR-Gs complex revealed the interaction interface of β2AR-Gs and structural changes upon complex formation [S. G. Rasmussen et al., Nature 477, 549–555 (2011)], yet, the dynamic process of the β2AR signaling through Gs and its preferential coupling to Gs over Gi is still not fully understood. Here, we utilize solution nuclear magnetic resonance (NMR) spectroscopy and supporting molecular dynamics (MD) simulations to monitor the conformational changes in the G protein coupling interface of the β2AR in response to the full agonist BI-167107 and Gs and Gi1. These results show that BI-167107 stabilizes conformational changes in four transmembrane segments (TM4, TM5, TM6, and TM7) prior to coupling to a G protein, and that the agonist-bound receptor conformation is different from the G protein coupled state. While most of the conformational changes observed in the β2AR are qualitatively the same for Gs and Gi1, we detected distinct differences between the β2AR-Gs and the β2AR-Gi1 complex in intracellular loop 2 (ICL2). Interactions with ICL2 are essential for activation of Gs. These differences between the β2AR-Gs and β2AR-Gi1 complexes in ICL2 may be key determinants for G protein coupling selectivity.

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