scholarly journals Molecular basis for high-affinity agonist binding in GPCRs

Science ◽  
2019 ◽  
Vol 364 (6442) ◽  
pp. 775-778 ◽  
Author(s):  
Tony Warne ◽  
Patricia C. Edwards ◽  
Andrew S. Doré ◽  
Andrew G. W. Leslie ◽  
Christopher G. Tate
Keyword(s):  
Hydrogen Bonds ◽  
G Protein ◽  
Molecular Basis ◽  
G Protein Coupled Receptors ◽  
Active State ◽  
Agonist Binding ◽  
Inactive State ◽  
High Affinity ◽  
Wide Range ◽  
G Protein Coupled

G protein–coupled receptors (GPCRs) in the G protein–coupled active state have higher affinity for agonists as compared with when they are in the inactive state, but the molecular basis for this is unclear. We have determined four active-state structures of the β1-adrenoceptor (β1AR) bound to conformation-specific nanobodies in the presence of agonists of varying efficacy. Comparison with inactive-state structures of β1AR bound to the identical ligands showed a 24 to 42% reduction in the volume of the orthosteric binding site. Potential hydrogen bonds were also shorter, and there was up to a 30% increase in the number of atomic contacts between the receptor and ligand. This explains the increase in agonist affinity of GPCRs in the active state for a wide range of structurally distinct agonists.

scholarly journals Molecular basis for high affinity agonist binding in GPCRs

2018 ◽  
Author(s):  
Tony Warne ◽  
Patricia C. Edwards ◽  
Andrew S. Doré ◽  
Andrew G. W. Leslie ◽  
Christopher G. Tate
Keyword(s):  
Hydrogen Bonds ◽  
Binding Site ◽  
G Protein ◽  
Molecular Basis ◽  
Agonist Binding ◽  
High Affinity ◽  
Protein Ligand Interactions ◽  
Wide Range ◽  
Ligand Interactions ◽  
G Protein Coupled

AbstractA characteristic of GPCRs in the G protein-coupled state is that the affinity of the agonist often increases significantly, but the molecular basis for this is unclear. We have determined six active-state structures of the β1-adrenoceptor (β1AR) bound to conformation-specific nanobodies in the presence of agonists of varying efficacy. A direct comparison with structures of β1AR in inactive states bound to the identical ligands showed a 24-42% reduction in the volume of the orthosteric binding site. Potential hydrogen bonds were also shorter, and there was up to a 30% increase in the number of atomic contacts between the receptor and ligand. GPCRs are highly conserved, so these factors will likely be essential in increasing the affinity of a wide range of structurally distinct agonists.One Sentence SummaryHigh affinity agonist binding to G protein-coupled GPCRs results from an increase in the number and strength of protein-ligand interactions.

scholarly journals Mini-review: antibody therapeutics targeting G protein-coupled receptors and ion channels

2020 ◽  
Vol 3 (4) ◽  
pp. 257-264
Author(s):  
Catherine J Hutchings
Keyword(s):  
Ion Channels ◽  
Research And Development ◽  
Small Molecules ◽  
G Protein ◽  
Clinical Studies ◽  
G Protein Coupled Receptors ◽  
Protein Targets ◽  
Antibody Therapeutics ◽  
Wide Range ◽  
G Protein Coupled

Abstract Antibodies are now well established as therapeutics with many additional advantages over small molecules and peptides relative to their selectivity, bioavailability, half-life and effector function. Major classes of membrane-associated protein targets include G protein-coupled receptors (GPCRs) and ion channels that are linked to a wide range of disease indications across all therapeutic areas. This mini-review summarizes the antibody target landscape for both GPCRs and ion channels as well as current progress in the respective research and development pipelines with some example case studies highlighted from clinical studies, including those being evaluated for the treatment of symptoms in COVID-19 infection.

scholarly journals Assembly of G Protein-Coupled Receptors onto Nanosized Bacterial Magnetic Particles Using Mms16 as an Anchor Molecule

2004 ◽  
Vol 70 (5) ◽  
pp. 2880-2885 ◽  
Author(s):  
Tomoko Yoshino ◽  
Masayoshi Takahashi ◽  
Haruko Takeyama ◽  
Yoshiko Okamura ◽  
Fukuichi Kato ◽  
...  
Keyword(s):  
G Protein ◽  
Magnetic Particles ◽  
Lipid Membrane ◽  
Specific Protein ◽  
G Protein Coupled Receptors ◽  
Native Conformation ◽  
Wide Range ◽  
Bacterial Magnetic Particles ◽  
Magnetospirillum Magneticum ◽  
G Protein Coupled

ABSTRACT G protein-coupled receptors (GPCRs) play a central role in a wide range of biological processes and are prime targets for drug discovery. GPCRs have large hydrophobic domains, and therefore purification of GPCRs from cells is frequently time-consuming and typically results in loss of native conformation. In this work, GPCRs have been successfully assembled into the lipid membrane of nanosized bacterial magnetic particles (BMPs) produced by the magnetic bacterium Magnetospirillum magneticum AMB-1. A BMP-specific protein, Mms16, was used as an anchor molecule, and localization of heterologous Mms16 on BMPs was confirmed by luciferase fusion studies. Stable luminescence was obtained from BMPs bearing Mms16 fused with luciferase at the C-terminal region. D1 dopamine receptor (D1R), a GPCR, was also efficiently assembled onto BMPs by using Mms16 as an anchor molecule. D1R-BMP complexes were simply extracted by magnetic separation from ruptured AMB-1 transformants. After washing, the complexes were ready to use for analysis. This system conveniently refines the native conformation of GPCRs without the need for detergent solubilization, purification, and reconstitution after cell disruption.

scholarly journals The molecular basis of subtype selectivity of human kinin G-protein-coupled receptors

2018 ◽  
Vol 14 (3) ◽  
pp. 284-290 ◽  
Author(s):  
Lisa Joedicke ◽  
Jiafei Mao ◽  
Georg Kuenze ◽  
Christoph Reinhart ◽  
Tejaswi Kalavacherla ◽  
...  
Keyword(s):  
G Protein ◽  
Molecular Basis ◽  
G Protein Coupled Receptors ◽  
Subtype Selectivity ◽  
G Protein Coupled

scholarly journals Specific oxylipins enhance vertebrate hematopoiesis via the receptor GPR132

2018 ◽  
Vol 115 (37) ◽  
pp. 9252-9257 ◽  
Author(s):  
Jamie L. Lahvic ◽  
Michelle Ammerman ◽  
Pulin Li ◽  
Megan C. Blair ◽  
Emma R. Stillman ◽  
...  
Keyword(s):  
Fatty Acids ◽  
G Protein ◽  
Cell Lines ◽  
Saturated Fatty Acids ◽  
Receptor Activation ◽  
G Protein Coupled Receptors ◽  
Hydroxy Fatty Acids ◽  
High Affinity ◽  
G Protein Coupled

Epoxyeicosatrienoic acids (EETs) are lipid-derived signaling molecules with cardioprotective and vasodilatory actions. We recently showed that 11,12-EET enhances hematopoietic induction and engraftment in mice and zebrafish. EETs are known to signal via G protein-coupled receptors, with evidence supporting the existence of a specific high-affinity receptor. Identification of a hematopoietic-specific EET receptor would enable genetic interrogation of EET signaling pathways, and perhaps clinical use of this molecule. We developed a bioinformatic approach to identify an EET receptor based on the expression of G protein-coupled receptors in cell lines with differential responses to EETs. We found 10 candidate EET receptors that are expressed in three EET-responsive cell lines, but not expressed in an EET-unresponsive line. Of these, only recombinant GPR132 showed EET-responsiveness in vitro, using a luminescence-based β-arrestin recruitment assay. Knockdown of zebrafish gpr132b prevented EET-induced hematopoiesis, and marrow from GPR132 knockout mice showed decreased long-term engraftment capability. In contrast to high-affinity EET receptors, GPR132 is reported to respond to additional hydroxy-fatty acids in vitro, and we found that these same hydroxy-fatty acids enhance hematopoiesis in the zebrafish. We conducted structure–activity relationship analyses using both cell culture and zebrafish assays on diverse medium-chain fatty acids. Certain oxygenated, unsaturated free fatty acids showed high activation of GPR132, whereas unoxygenated or saturated fatty acids had lower activity. Absence of the carbon-1 position carboxylic acid prevented activity, suggesting that this moiety is required for receptor activation. GPR132 responds to a select panel of oxygenated polyunsaturated fatty acids to enhance both embryonic and adult hematopoiesis.

scholarly journals Methods used to study the oligomeric structure of G-protein-coupled receptors

2017 ◽  
Vol 37 (2) ◽  
Author(s):  
Hui Guo ◽  
Su An ◽  
Richard Ward ◽  
Yang Yang ◽  
Ying Liu ◽  
...  
Keyword(s):  
Energy Transfer ◽  
G Protein ◽  
Resonance Energy Transfer ◽  
Resonance Energy ◽  
G Protein Coupled Receptors ◽  
Experimental Techniques ◽  
Distribution Analysis ◽  
Wide Range ◽  
And Function ◽  
G Protein Coupled

G-protein-coupled receptors (GPCRs), which constitute the largest family of cell surface receptors, were originally thought to function as monomers, but are now recognized as being able to act in a wide range of oligomeric states and indeed, it is known that the oligomerization state of a GPCR can modulate its pharmacology and function. A number of experimental techniques have been devised to study GPCR oligomerization including those based upon traditional biochemistry such as blue-native PAGE (BN-PAGE), co-immunoprecipitation (Co-IP) and protein-fragment complementation assays (PCAs), those based upon resonance energy transfer, FRET, time-resolved FRET (TR-FRET), FRET spectrometry and bioluminescence resonance energy transfer (BRET). Those based upon microscopy such as FRAP, total internal reflection fluorescence microscopy (TIRFM), spatial intensity distribution analysis (SpIDA) and various single molecule imaging techniques. Finally with the solution of a growing number of crystal structures, X-ray crystallography must be acknowledged as an important source of discovery in this field. A different, but in many ways complementary approach to the use of more traditional experimental techniques, are those involving computational methods that possess obvious merit in the study of the dynamics of oligomer formation and function. Here, we summarize the latest developments that have been made in the methods used to study GPCR oligomerization and give an overview of their application.

Molecular basis of G protein-coupled receptor high affinity for gastrin-releasing peptide (GRP)

2003 ◽  
Vol 124 (4) ◽  
pp. A469
Author(s):  
Tomoo Nakagawa ◽  
Jose A. Tapia ◽  
Kenji Tokita ◽  
Samuel Mantey ◽  
Michael Schumann ◽  
...  
Keyword(s):  
G Protein ◽  
Molecular Basis ◽  
G Protein Coupled Receptor ◽  
Gastrin Releasing Peptide ◽  
High Affinity ◽  
G Protein Coupled

scholarly journals Molecular Basis of Secretin Docking to Its Intact Receptor Using Multiple Photolabile Probes Distributed throughout the Pharmacophore

2011 ◽  
Vol 286 (27) ◽  
pp. 23888-23899 ◽  
Author(s):  
Maoqing Dong ◽  
Polo C.-H. Lam ◽  
Delia I. Pinon ◽  
Keiko Hosohata ◽  
Andrew Orry ◽  
...  
Keyword(s):  
G Protein ◽  
Molecular Basis ◽  
Peptide Mapping ◽  
G Protein Coupled Receptors ◽  
Peptide Ligands ◽  
Amino Terminus ◽  
Amino Terminal ◽  
Binding Cleft ◽  
Mutant Receptors ◽  
G Protein Coupled

The molecular basis of ligand binding and activation of family B G protein-coupled receptors is not yet clear due to the lack of insight into the structure of intact receptors. Although NMR and crystal structures of amino-terminal domains of several family members support consistency in general structural motifs that include a peptide-binding cleft, there are variations in the details of docking of the carboxyl terminus of peptide ligands within this cleft, and there is no information about siting of the amino terminus of these peptides. There are also no empirical data to orient the receptor amino terminus relative to the core helical bundle domain. Here, we prepared a series of five new probes, incorporating photolabile moieties into positions 2, 15, 20, 24, and 25 of full agonist secretin analogues. Each bound specifically to the receptor and covalently labeled single distinct receptor residues. Peptide mapping of labeled wild-type and mutant receptors identified that the position 15, 20, and 25 probes labeled residues within the distal amino terminus of the receptor, whereas the position 24 probe labeled the amino terminus adjacent to TM1. Of note, the position 2 probe labeled a residue within the first extracellular loop of the receptor, a region not previously labeled, providing an important new constraint for docking the amino-terminal region of secretin to its receptor core. These additional experimentally derived constraints help to refine our understanding of the structure of the secretin-intact receptor complex and provide new insights into understanding the molecular mechanism for activation of family B G protein-coupled receptors.

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