scholarly journals Structural Requirements for the Activation of the Human Growth Hormone Secretagogue Receptor by Peptide and Nonpeptide Secretagogues

1998 ◽  
Vol 12 (1) ◽  
pp. 137-145 ◽  
Author(s):  
Scott D. Feighner ◽  
Andrew D. Howard ◽  
Kristine Prendergast ◽  
Oksana C. Palyha ◽  
Donna L. Hreniuk ◽  
...  
Keyword(s):  
Ligand Binding ◽  
Binding Site ◽  
G Protein ◽  
Human Growth ◽  
G Protein Coupled Receptors ◽  
Site Directed Mutagenesis ◽  
Agonist Binding ◽  
Growth Hormone Secretagogue Receptor ◽  
Growth Hormone Secretagogue ◽  
G Protein Coupled

Abstract Antibodies raised against an intracellular and extracellular domain of the GH secretagogue receptor (GHS-R) confirmed that its topological orientation in the lipid bilayer is as predicted for G protein-coupled receptors with seven transmembrane domains. A strategy for mapping the agonist-binding site of the human GHS-R was conceived based on our understanding of ligand binding in biogenic amine and peptide hormone G protein-coupled receptors. Using site-directed mutagenesis and molecular modeling, we classified GHS peptide and nonpeptide agonist binding in the context of its receptor environment. All peptide and nonpeptide ligand classes shared a common binding domain in transmembrane (TM) region 3 of the GHS-R. This finding was based on TM-3 mutation E124Q, which eliminated the counter-ion to the shared basic N+ group of all GHSs and resulted in a nonfunctional receptor. Restoration of function for the E124Q mutant was achieved by a complementary change in the MK-0677 ligand through modification of its amine side-chain to the corresponding alcohol. Contacts in other TM domains [TM-2 (D99N), TM-5 (M213K, S117A), TM-6 (H280F), and extracellular loop 1 (C116A)] of the receptor revealed specificity for the different peptide, benzolactam, and spiroindolane GHSs. GHS-R agonism, therefore, does not require identical disposition of all agonist classes at the ligand-binding site. Our results support the hypothesis that the ligand-binding pocket in the GHS-R is spatially disposed similarly to the well characterized catechol-binding site in theβ 2-adrenergic receptor.

scholarly journals G Protein-Coupled Receptors in Taste Physiology and Pharmacology

2020 ◽  
Vol 11 ◽  
Author(s):  
Raise Ahmad ◽  
Julie E. Dalziel
Keyword(s):  
Binding Site ◽  
G Protein ◽  
Molecular Mechanisms ◽  
Taste Receptor ◽  
Receptor Activation ◽  
G Protein Coupled Receptors ◽  
Type I ◽  
Taste Receptors ◽  
Agonist Binding ◽  
G Protein Coupled

Heterotrimeric G protein-coupled receptors (GPCRs) comprise the largest receptor family in mammals and are responsible for the regulation of most physiological functions. Besides mediating the sensory modalities of olfaction and vision, GPCRs also transduce signals for three basic taste qualities of sweet, umami (savory taste), and bitter, as well as the flavor sensation kokumi. Taste GPCRs reside in specialised taste receptor cells (TRCs) within taste buds. Type I taste GPCRs (TAS1R) form heterodimeric complexes that function as sweet (TAS1R2/TAS1R3) or umami (TAS1R1/TAS1R3) taste receptors, whereas Type II are monomeric bitter taste receptors or kokumi/calcium-sensing receptors. Sweet, umami and kokumi receptors share structural similarities in containing multiple agonist binding sites with pronounced selectivity while most bitter receptors contain a single binding site that is broadly tuned to a diverse array of bitter ligands in a non-selective manner. Tastant binding to the receptor activates downstream secondary messenger pathways leading to depolarization and increased intracellular calcium in TRCs, that in turn innervate the gustatory cortex in the brain. Despite recent advances in our understanding of the relationship between agonist binding and the conformational changes required for receptor activation, several major challenges and questions remain in taste GPCR biology that are discussed in the present review. In recent years, intensive integrative approaches combining heterologous expression, mutagenesis and homology modeling have together provided insight regarding agonist binding site locations and molecular mechanisms of orthosteric and allosteric modulation. In addition, studies based on transgenic mice, utilizing either global or conditional knock out strategies have provided insights to taste receptor signal transduction mechanisms and their roles in physiology. However, the need for more functional studies in a physiological context is apparent and would be enhanced by a crystallized structure of taste receptors for a more complete picture of their pharmacological mechanisms.

scholarly journals Exploring a new ligand binding site of G protein-coupled receptors

2018 ◽  
Vol 9 (31) ◽  
pp. 6480-6489 ◽  
Author(s):  
H. C. Stephen Chan ◽  
Jingjing Wang ◽  
Krzysztof Palczewski ◽  
Slawomir Filipek ◽  
Horst Vogel ◽  
...  
Keyword(s):  
Ligand Binding ◽  
Binding Site ◽  
G Protein ◽  
Md Simulations ◽  
Binding Pocket ◽  
G Protein Coupled Receptors ◽  
Ligand Binding Site ◽  
Endogenous Ligand ◽  
G Protein Coupled

A new binding pocket of the endogenous ligand has been discovered by MD simulations.

Multiparametric Homogeneous Method for Identification of Ligand Binding to G Protein-Coupled Receptors: Receptor–Ligand Binding and β-Arrestin Assay

2013 ◽  
Vol 85 (4) ◽  
pp. 2276-2281 ◽  
Author(s):  
Kari Kopra ◽  
Markus Kainulainen ◽  
Piia Mikkonen ◽  
Anita Rozwandowicz-Jansen ◽  
Pekka Hänninen ◽  
...  
Keyword(s):  
Ligand Binding ◽  
G Protein ◽  
G Protein Coupled Receptors ◽  
Receptor Ligand ◽  
Homogeneous Method ◽  
G Protein Coupled

Family Resemblances? Ligand Binding and Activation of Family A and B G-Protein-Coupled Receptors

2007 ◽  
Vol 35 (4) ◽  
pp. 707-708 ◽  
Author(s):  
D.R. Poyner ◽  
M. Wheatley
Keyword(s):  
Ligand Binding ◽  
G Protein ◽  
Present Article ◽  
G Protein Coupled Receptors ◽  
Society Meeting ◽  
Family Resemblances ◽  
G Protein Coupled ◽  
Compare And Contrast ◽  
Present Meeting

In April 2007, the Biochemical Society held a meeting to compare and contrast ligand binding and activation of Family A and B GPCRs (G-protein-coupled receptors). Being the largest class, Family A GPCRs usually receive the most attention, although a previous Biochemical Society meeting has focused on Family B GPCRs. The aim of the present meeting was to bring researchers of both families together in order to identify commonalities between the two. The present article introduces the proceedings of the meeting, briefly commenting on the focus of each of the following articles.

scholarly journals Locating ligand binding sites in G-protein coupled receptors using combined information from docking and sequence conservation

2018 ◽  
Author(s):  
Ashley R. Vidad ◽  
Stephen Macaspac ◽  
Ho-Leung Ng
Keyword(s):  
Ligand Binding ◽  
G Protein ◽  
Binding Sites ◽  
G Protein Coupled Receptors ◽  
Sequence Conservation ◽  
Surface Geometry ◽  
A2a Adenosine Receptors ◽  
Ligand Binding Sites ◽  
G Protein Coupled ◽  
Homology Models

AbstractG-protein coupled receptors (GPCRs) are the largest protein family of drug targets. Detailed mechanisms of binding are unknown for many important GPCR-ligand pairs due to the difficulties of GPCR recombinant expression, biochemistry, and crystallography. We describe our new method, ConDock, for predicting ligand binding sites in GPCRs using combined information from surface conservation and docking starting from crystal structures or homology models. We demonstrate the effectiveness of ConDock on well-characterized GPCRs such as the β2 adrenergic and A2A adenosine receptors. We also demonstrate that ConDock successfully predicts ligand binding sites from high-quality homology models. Finally, we apply ConDock to predict ligand binding sites on a structurally uncharacterized GPCR, GPER. GPER is the G-protein coupled estrogen receptor, with four known ligands: estradiol, G1, G15, and tamoxifen. ConDock predicts that all four ligands bind to the same location on GPER, centered on L119, H307, and N310; this site is deeper in the receptor cleft than predicted by previous studies. We compare the sites predicted by ConDock and traditional methods that utilize information from surface geometry, surface conservation, and ligand chemical interactions. Incorporating sequence conservation information in ConDock overcomes errors introduced from physics-based scoring functions and homology modeling.

scholarly journals Probing the correlation between ligand efficacy and conformational diversity at the α1A-adrenoreceptor reveals allosteric coupling of its microswitches

2020 ◽  
Vol 295 (21) ◽  
pp. 7404-7417 ◽  
Author(s):  
Feng-Jie Wu ◽  
Lisa M. Williams ◽  
Alaa Abdul-Ridha ◽  
Avanka Gunatilaka ◽  
Tasneem M. Vaid ◽  
...  
Keyword(s):  
Ligand Binding ◽  
Binding Site ◽  
G Protein ◽  
Conformational Dynamics ◽  
Smooth Muscle Contraction ◽  
Allosteric Coupling ◽  
Gpcr Activation ◽  
Methionine Residues ◽  
G Protein Coupled ◽  
Receptor Conformation

G protein–coupled receptors (GPCRs) use a series of conserved microswitches to transmit signals across the cell membrane via an allosteric network encompassing the ligand-binding site and the G protein-binding site. Crystal structures of GPCRs provide snapshots of their inactive and active states, but poorly describe the conformational dynamics of the allosteric network that underlies GPCR activation. Here, we analyzed the correlation between ligand binding and receptor conformation of the α1A-adrenoreceptor, a GPCR that stimulates smooth muscle contraction in response to binding noradrenaline. NMR of [13CϵH3]methionine-labeled α1A-adrenoreceptor variants, each exhibiting differing signaling capacities, revealed how different classes of ligands modulate the conformational equilibria of this receptor. [13CϵH3]Methionine residues near the microswitches exhibited distinct states that correlated with ligand efficacies, supporting a conformational selection mechanism. We propose that allosteric coupling among the microswitches controls the conformation of the α1A-adrenoreceptor and underlies the mechanism of ligand modulation of GPCR signaling in cells.

New small molecule fluorescent probes for G protein-coupled receptors: valuable tools for drug discovery

2021 ◽  
Vol 13 (1) ◽  
pp. 63-90
Author(s):  
Joshua W Conner ◽  
Daniel P Poole ◽  
Manuela Jörg ◽  
Nicholas A Veldhuis
Keyword(s):  
Drug Discovery ◽  
Ligand Binding ◽  
G Protein ◽  
Small Molecule ◽  
G Protein Coupled Receptors ◽  
Signaling Proteins ◽  
Drug Candidates ◽  
Compound Screening ◽  
G Protein Coupled ◽  
Single Approach

G protein-coupled receptors (GPCRs) are essential signaling proteins and tractable therapeutic targets. To develop new drug candidates, GPCR drug discovery programs require versatile, sensitive pharmacological tools for ligand binding and compound screening. With the availability of new imaging modalities and proximity-based ligand binding technologies, fluorescent ligands offer many advantages and are increasingly being used, yet labeling small molecules remains considerably more challenging relative to peptides. Focusing on recent fluorescent small molecule studies for family A GPCRs, this review addresses some of the key challenges, synthesis approaches and structure–activity relationship considerations, and discusses advantages of using high-resolution GPCR structures to inform conjugation strategies. While no single approach guarantees successful labeling without loss of affinity or selectivity, the choice of fluorophore, linker type and site of attachment have proved to be critical factors that can significantly affect their utility in drug discovery programs, and as discussed, can sometimes lead to very unexpected results.

scholarly journals A Microdomain Formed by the Extracellular Ends of the Transmembrane Domains Promotes Activation of the G Protein-Coupled α-Factor Receptor

2004 ◽  
Vol 24 (5) ◽  
pp. 2041-2051 ◽  
Author(s):  
Jennifer C. Lin ◽  
Ken Duell ◽  
James B. Konopka
Keyword(s):  
Ligand Binding ◽  
G Protein ◽  
Transmembrane Domain ◽  
Receptor Activation ◽  
G Protein Coupled Receptors ◽  
Transmembrane Domains ◽  
Consecutive Series ◽  
Subsequent Step ◽  
G Protein Coupled ◽  
Specific Reagent

ABSTRACT The α-factor receptor (Ste2p) that promotes mating in Saccharomyces cerevisiae is similar to other G protein-coupled receptors (GPCRs) in that it contains seven transmembrane domains. Previous studies suggested that the extracellular ends of the transmembrane domains are important for Ste2p function, so a systematic scanning mutagenesis was carried out in which 46 residues near the ends of transmembrane domains 1, 2, 3, 4, and 7 were replaced with cysteine. These mutants complement mutations constructed previously near the ends of transmembrane domains 5 and 6 to analyze all the extracellular ends. Eight new mutants created in this study were partially defective in signaling (V45C, N46C, T50C, A52C, L102C, N105C, L277C, and A281C). Treatment with 2-([biotinoyl] amino) ethyl methanethiosulfonate, a thiol-specific reagent that reacts with accessible cysteine residues but not membrane-embedded cysteines, identified a drop in the level of reactivity over a consecutive series of residues that was inferred to be the membrane boundary. An unusual prolonged zone of intermediate reactivity near the extracellular end of transmembrane domain 2 suggests that this region may adopt a special structure. Interestingly, residues implicated in ligand binding were mainly accessible, whereas residues involved in the subsequent step of promoting receptor activation were mainly inaccessible. These results define a receptor microdomain that provides an important framework for interpreting the mechanisms by which functionally important residues contribute to ligand binding and activation of Ste2p and other GPCRs.

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