Structure and dynamics of G protein-coupled receptor–bound ghrelin reveal the critical role of the octanoyl chain

Ghrelin plays a central role in controlling major biological processes. As for other G protein-coupled receptor (GPCR) peptide agonists, the structure and dynamics of ghrelin bound to its receptor remain obscure. Using a combination of solution-state NMR and molecular modeling, we demonstrate that binding to the growth hormone secretagogue receptor is accompanied by a conformational change in ghrelin that structures its central region, involving the formation of a well-defined hydrophobic core. By comparing its acylated and nonacylated forms, we conclude that the ghrelin octanoyl chain is essential to form the hydrophobic core and promote access of ghrelin to the receptor ligand-binding pocket. The combination of coarse-grained molecular dynamics studies and NMR should prove useful in improving our mechanistic understanding of the complex conformational space explored by a natural peptide agonist when binding to its GPCR. Such information should also facilitate the design of new ghrelin receptor-selective drugs.

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G protein-coupled receptor 120 signaling regulates ghrelin secretion in vivo and in vitro

AJP Endocrinology and Metabolism ◽

10.1152/ajpendo.00306.2013 ◽

2014 ◽

Vol 306 (1) ◽

pp. E28-E35 ◽

Cited By ~ 54

Author(s):

Zhi Gong ◽

Makoto Yoshimura ◽

Sayaka Aizawa ◽

Reiko Kurotani ◽

Jeffrey M. Zigman ◽

...

Keyword(s):

G Protein ◽

Cell Lines ◽

G Protein Coupled Receptor ◽

Growth Hormone Secretagogue Receptor ◽

Growth Hormone Secretagogue ◽

Expression Levels ◽

Ghrelin Secretion ◽

G Protein Coupled

Ghrelin, an endogenous ligand for the growth hormone secretagogue receptor, is produced predominantly in the stomach. It has been reported that endogenous ghrelin levels are increased by fasting and decreased immediately after feeding and that fasting-induced ghrelin release is controlled by the sympathetic nervous system. However, the mechanisms of plasma ghrelin decrement after feeding are poorly understood. Here, we studied the control of ghrelin secretion using ghrelin-producing cell lines and found that these cells express high levels of mRNA encoding G-protein coupled receptor 120 (GPR120). Addition of GW-9508 (a GPR120 chemical agonist) and α-linolenic acid (a natural ligand for GPR120) inhibited the secretion of ghrelin by ∼50 and 70%, respectively. However, the expression levels of preproghrelin and ghrelin O-acyltransferase (GOAT) mRNAs were not influenced by GW-9508. In contrast, the expression levels of prohormone convertase 1 were decreased significantly by GW-9508 incubation. Moreover, we observed that the inhibitory effect of GW-9508 on ghrelin secretion was blocked by a small interfering RNA (siRNA) targeting the sequence of GPR120. Furthermore, pretreatment with GW-9508 blocked the effect of the norepinephrine (NE)-induced ghrelin elevation in ghrelin cell lines. In addition, we showed that GW-9508 inhibited ghrelin secretion via extracellular signal-regulated kinase activity in ghrelin cell lines. Finally, we found that GW-9508 decreased plasma ghrelin levels in mice. These results suggest that the decrease of ghrelin secretion after feeding is induced partially by long-chain fatty acids that act directly on gastric GPR120-expressing ghrelin cells.

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An Efficient Strategy to Estimate Thermodynamics and Kinetics of G Protein-Coupled Receptor Activation Using Metadynamics and Maximum Caliber

10.1101/367888 ◽

2018 ◽

Author(s):

Derya Meral ◽

Davide Provasi ◽

Marta Filizola

Keyword(s):

G Protein ◽

Adaptive Sampling ◽

Receptor Activation ◽

Conformational Space ◽

Kinetic Properties ◽

G Protein Coupled Receptor ◽

Efficient Strategy ◽

Gpcr Activation ◽

Kinetic Rates ◽

G Protein Coupled

ABSTRACTComputational strategies aimed at unveiling the thermodynamic and kinetic properties of G Protein-Coupled Receptor (GPCR) activation require extensive molecular dynamics simulations of the receptor embedded in an explicit lipid-water environment. A possible method for efficiently sampling the conformational space of such a complex system is metadynamics (MetaD) with path collective variables (CV). Here, we applied well-tempered MetaD with path CVs to one of the few GPCRs for which both inactive and fully active experimental structures are available, the μ-opioid receptor (MOR), and assessed the ability of this enhanced sampling method to estimate thermodynamic properties of receptor activation in line with those obtained by more computationally expensive adaptive sampling protocols. While n-body information theory (nBIT) analysis of these simulations confirmed that MetaD can efficiently characterize ligand-induced allosteric communication across the receptor, standard MetaD cannot be used directly to derive kinetic rates because transitions are accelerated by a bias potential. Applying the principle of Maximum Caliber (MaxCal) to the free-energy landscape of morphine-bound MOR reconstructed from MetaD, we obtained Markov State Models (MSMs) that yield kinetic rates of MOR activation in agreement with those obtained by adaptive sampling. Taken together, these results suggest that the MetaD-MaxCal combination creates an efficient strategy for estimating thermodynamic and kinetic properties of GPCR activation at an affordable computational cost.

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Hybrid MM/CG Webserver: Automatic Set Up of Molecular Mechanics/Coarse-Grained Simulations for Human G Protein-Coupled Receptor/Ligand Complexes

Frontiers in Molecular Biosciences ◽

10.3389/fmolb.2020.576689 ◽

2020 ◽

Vol 7 ◽

Cited By ~ 2

Author(s):

Jakob Schneider ◽

Rui Ribeiro ◽

Mercedes Alfonso-Prieto ◽

Paolo Carloni ◽

Alejandro Giorgetti

Keyword(s):

Molecular Mechanics ◽

G Protein ◽

Coarse Grained ◽

G Protein Coupled Receptor ◽

Receptor Ligand ◽

Coarse Grained Simulations ◽

Set Up ◽

G Protein Coupled

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Inhibition of Ebola and Marburg Virus Entry by G Protein-Coupled Receptor Antagonists

Journal of Virology ◽

10.1128/jvi.01337-15 ◽

2015 ◽

Vol 89 (19) ◽

pp. 9932-9938 ◽

Cited By ~ 49

Author(s):

Han Cheng ◽

Calli M. Lear-Rooney ◽

Lisa Johansen ◽

Elizabeth Varhegyi ◽

Zheng W. Chen ◽

...

Keyword(s):

Antiviral Activity ◽

G Protein ◽

High Mortality ◽

Ebola Virus ◽

Critical Role ◽

Marburg Virus ◽

Severe Illness ◽

G Protein Coupled Receptor ◽

Chemical Library ◽

G Protein Coupled

ABSTRACTFiloviruses, consisting of Ebola virus (EBOV) and Marburg virus (MARV), are among the most lethal infectious threats to mankind. Infections by these viruses can cause severe hemorrhagic fevers in humans and nonhuman primates with high mortality rates. Since there is currently no vaccine or antiviral therapy approved for humans, there is an urgent need to develop prophylactic and therapeutic options for use during filoviral outbreaks and bioterrorist attacks. One of the ideal targets against filoviral infection and diseases is at the entry step, which is mediated by the filoviral glycoprotein (GP). In this report, we screened a chemical library of small molecules and identified numerous inhibitors, which are known G protein-coupled receptor (GPCR) antagonists targeting different GPCRs, including histamine receptors, 5-HT (serotonin) receptors, muscarinic acetylcholine receptor, and adrenergic receptor. These inhibitors can effectively block replication of both infectious EBOV and MARV, indicating a broad antiviral activity of the GPCR antagonists. The time-of-addition experiment and microscopic studies suggest that GPCR antagonists block filoviral entry at a step following the initial attachment but prior to viral/cell membrane fusion. These results strongly suggest that GPCRs play a critical role in filoviral entry and GPCR antagonists can be developed as an effective anti-EBOV/MARV therapy.IMPORTANCEInfection of Ebola virus and Marburg virus can cause severe illness in humans with a high mortality rate, and currently there is no FDA-approved vaccine or therapeutic treatment available. The 2013-2015 epidemic in West Africa underscores a lack of our understanding in the infection and pathogenesis of these viruses and the urgency of drug discovery and development. In this study, we have identified numerous inhibitors that are known G protein-coupled receptor (GPCR) antagonists targeting different GPCRs. These inhibitors can effectively block replication of both infectious EBOV and MARV, indicating a broad antiviral activity of the GPCR antagonists. Our results strongly suggest that GPCRs play a critical role in filoviral entry and GPCR antagonists can be developed as an effective anti-EBOV/MARV therapy.

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Concerted conformational dynamics and water movements in the ghrelin G protein-coupled receptor

eLife ◽

10.7554/elife.63201 ◽

2021 ◽

Vol 10 ◽

Author(s):

Maxime Louet ◽

Marina Casiraghi ◽

Marjorie Damian ◽

Mauricio GS Costa ◽

Pedro Renault ◽

...

Keyword(s):

G Protein ◽

Unnatural Amino Acids ◽

Conformational Dynamics ◽

Signal Propagation ◽

Water Molecules ◽

Growth Hormone Secretagogue Receptor ◽

Growth Hormone Secretagogue ◽

Ghrelin Receptor ◽

Tightly Coupled ◽

G Protein Coupled

There is increasing support for water molecules playing a role in signal propagation through G protein-coupled receptors (GPCRs). However, exploration of the hydration features of GPCRs is still in its infancy. Here, we combined site-specific labeling with unnatural amino acids to molecular dynamics to delineate how local hydration of the ghrelin receptor growth hormone secretagogue receptor (GHSR) is rearranged upon activation. We found that GHSR is characterized by a specific hydration pattern that is selectively remodeled by pharmacologically distinct ligands and by the lipid environment. This process is directly related to the concerted movements of the transmembrane domains of the receptor. These results demonstrate that the conformational dynamics of GHSR are tightly coupled to the movements of internal water molecules, further enhancing our understanding of the molecular bases of GPCR-mediated signaling.

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Acute loss of Cell–Cell Communication Caused by G Protein–coupled Receptors: A Critical Role for c-Src

The Journal of Cell Biology ◽

10.1083/jcb.140.5.1199 ◽

1998 ◽

Vol 140 (5) ◽

pp. 1199-1209 ◽

Cited By ~ 84

Author(s):

Friso R. Postma ◽

Trudi Hengeveld ◽

Jacqueline Alblas ◽

Ben N.G. Giepmans ◽

Gerben C.M. Zondag ◽

...

Keyword(s):

Protein Kinase ◽

Gap Junction ◽

G Protein ◽

Critical Role ◽

Cell Communication ◽

G Protein Coupled Receptors ◽

G Protein Coupled Receptor ◽

Junction Protein ◽

G Protein Coupled ◽

Cell Cell

Gap junctions mediate cell–cell communication in almost all tissues, but little is known about their regulation by physiological stimuli. Using a novel single-electrode technique, together with dye coupling studies, we show that in cells expressing gap junction protein connexin43, cell–cell communication is rapidly disrupted by G protein–coupled receptor agonists, notably lysophosphatidic acid, thrombin, and neuropeptides. In the continuous presence of agonist, junctional communication fully recovers within 1–2 h of receptor stimulation. In contrast, a desensitization-defective G protein–coupled receptor mediates prolonged uncoupling, indicating that recovery of communication is controlled, at least in part, by receptor desensitization. Agonist-induced gap junction closure consistently follows inositol lipid breakdown and membrane depolarization and coincides with Rho-mediated cytoskeletal remodeling. However, we find that gap junction closure is independent of Ca2+, protein kinase C, mitogen-activated protein kinase, or membrane potential, and requires neither Rho nor Ras activation. Gap junction closure is prevented by tyrphostins, by dominant-negative c-Src, and in Src-deficient cells. Thus, G protein–coupled receptors use a Src tyrosine kinase pathway to transiently inhibit connexin43-based cell–cell communication.

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