Properties of the beta 1- and beta 2-adrenergic receptor subtypes revealed by molecular cloning.

1989 ◽  
Vol 35 (5) ◽  
pp. 721-725 ◽  
Author(s):  
T Frielle ◽  
M G Caron ◽  
R J Lefkowitz
Keyword(s):  
Adrenergic Receptor ◽  
Radioligand Binding ◽  
Binding Pocket ◽  
Site Directed Mutagenesis ◽  
Xenopus Laevis Oocytes ◽  
Receptor Subtypes ◽  
Subtype Selectivity ◽  
Beta 2 ◽  
Transmembrane Regions ◽  
Chimeric Rna

Abstract The beta 1- and beta 2-adrenergic receptor subtypes are biochemically and functionally similar, because both receptors mediate the catecholamine-dependent activation of adenylate cyclase through the GTP-binding protein, Gs. Pharmacologically, the two receptors can be distinguished on the basis of their relative affinities for the agonists epinephrine and norepinephrine as well as their affinities for several selective antagonists. The primary structures of the human beta 1- and beta 2-adrenergic receptors have recently been deduced from the cloning of their genes and (or) cDNAs, revealing high sequence homology and a membrane topography of seven putative transmembrane regions similar to that of rhodopsin. Chimeric beta 1/beta 2-adrenergic receptor cDNAs have been constructed by site-directed mutagenesis and the chimeric RNA transcripts expressed in Xenopus laevis oocytes. The pharmacological properties of the expressed chimeric receptor proteins were assessed by radioligand binding utilizing subtype-selective agonists and antagonists. Apparently, several of the putative transmembrane regions contribute significantly to the determination of subtype selectivity, presumably by formation of a ligand-binding pocket, with determinants for agonist and antagonist binding being distinguishable.

scholarly journals Molecular Docking Study on the Binding Mode of Cardioselective Phenoxyaminopropanol Blocker into β-adrenergic Receptor Subtypes

2012 ◽  
Vol 59 (2) ◽  
pp. 44-53
Author(s):  
M. Polakovičová ◽  
R. Čižmáriková
Keyword(s):  
Molecular Docking ◽  
Adrenergic Receptor ◽  
Docking Study ◽  
Binding Pocket ◽  
Binding Mode ◽  
Receptor Subtypes ◽  
Molecular Docking Study ◽  
Subtype Selectivity ◽  
Extracellular Region ◽  
Selective Ligand

AbstractStructural understanding of subtype specific ligand-binding pocket variations and interactions of ligand with receptor may facilitate design of novel selective drugs. To gain insights into the subtype selectivity of β-blockers we performed flexible molecular docking study to analyze the interaction mode of cardioselective phenoxyaminopropanol blocker into the β1 and β2-adrenergic receptor. The binding site analysis reveals a strong identity between important amino acid residues and interactions with ligand in orthosteric catecholamine- binding pocket. The differences in the binding mode of selective ligand have been identified in the extracellular region of receptor subtypes.

Insights into the Structural Aspects of the mGlu Receptor Orthosteric Binding Site

2019 ◽  
Vol 19 (26) ◽  
pp. 2421-2446 ◽  
Author(s):  
Junliang Hao ◽  
Qi Chen
Keyword(s):  
Binding Pocket ◽  
Receptor Subtypes ◽  
X Ray ◽  
Mglu Receptor ◽  
Metabotropic Glutamate ◽  
Amino Terminal ◽  
Subtype Selectivity ◽  
Agonist And Antagonist ◽  
Amino Terminal Domain ◽  
Structural Aspects

The amino terminal domain (ATD) of the metabotropic glutamate (mGlu) receptors contains the orthosteric glutamate recognition site, which is highly conserved across the eight mGlu receptor subtypes. In total, 29 X-ray crystal structures of the mGlu ATD proteins have been reported to date. These structures span across 3 subgroups and 6 subtypes, and include apo, agonist- and antagonist-bound structures. We will discuss the insights gained from the analysis of these structures with the focus on the interactions contributing to the observed group and subtype selectivity for select agonists. Furthermore, we will define the full expanded orthosteric ligand binding pocket (LBP) of the mGlu receptors, and discuss the macroscopic features of the mGlu ATD proteins.

beta-Adrenergic receptors in the developing rabbit lung

1981 ◽  
Vol 240 (4) ◽  
pp. E351-E357 ◽  
Author(s):  
J. A. Whitsett ◽  
M. A. Manton ◽  
C. Darovec-Beckerman ◽  
K. G. Adams ◽  
J. J. Moore
Keyword(s):  
Adenylate Cyclase ◽  
Adrenergic Receptors ◽  
Adrenergic Receptor ◽  
Cyclase Activity ◽  
Adenylate Cyclase Activity ◽  
Receptor Subtypes ◽  
Beta Adrenergic Receptors ◽  
Rabbit Lung ◽  
Beta Adrenergic ◽  
Beta 2

beta-Adrenergic receptors and catecholamine-sensitive adenylate cyclase were identified and partially characterized in membrane fractions of rabbit lungs from day 25 of gestation to adulthood with the beta-adrenergic antagonists (--)-[3H]dihydroalprenolol [(--)-[3H]DHA] and (--)-[125I]iodohydroxybenzylpindolol [(--)-[125I]HYP]. beta-Adrenergic receptor number (Bmax) increased 11.5-fold during this time period, increasing progressively during the latter days of gestation and the early neonatal period, from 37 +/- 10 fmol/mg protein at 25 days gestation to 425 +/- 51 fmol/mg in the adult rabbit lung (mean +/- SD). Receptor affinity for (--)-[3H]DHA (KD = 1.8 nM) or (--)-[125I]HYP (KD - 0.104 nM) and the proportion of beta 1- and beta 2-adrenergic receptor subtypes (60% beta 1 and 40% beta 2) did not change with advancing age. Basal adenylate cyclase activity in lung homogenates decreased significantly with increasing age, whereas the activity in the presence of catecholamine or NaF remained nearly constant. Catecholamines stimulated adenylate cyclase activity at all ages studied supporting a role of the maturation of beta-adrenergic receptors in the regulation of pulmonary function.

scholarly journals Mutational Analysis of the α1a-Adrenergic Receptor Binding Pocket of Antagonists by Radioligand Binding Assay

2008 ◽  
Vol 31 (4) ◽  
pp. 598-601 ◽  
Author(s):  
Maruf Ahmed ◽  
Murad Hossain ◽  
Mohiuddin Ahmed Bhuiyan ◽  
Masaji Ishiguro ◽  
Takashi Tanaka ◽  
...  
Keyword(s):  
Receptor Binding ◽  
Adrenergic Receptor ◽  
Binding Assay ◽  
Mutational Analysis ◽  
Radioligand Binding ◽  
Binding Pocket ◽  
Radioligand Binding Assay

scholarly journals Genetic regulation of β2-adrenergic receptors in 3T3-L1 fibroblasts

1989 ◽  
Vol 260 (1) ◽  
pp. 53-59 ◽  
Author(s):  
M T Nakada ◽  
K M Haskell ◽  
D J Ecker ◽  
J M Stadel ◽  
S T Crooke
Keyword(s):  
Phorbol Ester ◽  
Adrenergic Receptors ◽  
Adrenergic Receptor ◽  
Genetic Regulation ◽  
Receptor Subtype ◽  
Receptor Subtypes ◽  
Regulatory Sequences ◽  
Beta Adrenergic Receptor ◽  
Beta Adrenergic ◽  
Beta 2

The beta 2-adrenergic receptor from mouse 3T3-L1 cells is up-regulated through genetic mechanisms by glucocorticoids and butyrate. To study the genetic regulation of these receptors, we sequenced a 5 kb region of genomic DNA from 3T3-L1 cells, containing the beta-adrenergic receptor gene and approx. 1.5 kb of both 5′ and 3′ flanking sequences. The sequence contained one copy of an 8 bp consensus sequence which can confer phorbol ester-responsiveness to genes. Phorbol esters attenuated the up-regulation of beta 2-adrenergic receptors by glucocorticoids but not by butyrate. This effect was probably due to a phorbol ester-induced decrease in glucocorticoid receptor number. Using methylation-sensitive restriction enzymes, we examined the methylation of a CG-rich region occurring 5′ to the gene and did not detect any changes in methylation of this region upon dexamethasone or butyrate treatment. A total of 16 putative glucocorticoid response elements were found which may mediate the glucocorticoid-induced increase in beta 2-adrenergic receptors. A comparison of the regulatory sequences of the two beta-adrenergic receptor subtypes from human and mouse confirms the observed physiological controls of receptor subtype expression and offers an explanation as to why the subtypes differ in genetic regulation.

Characterization, expression, and hormonal control of a thymic beta 2-adrenergic receptor

1994 ◽  
Vol 267 (5) ◽  
pp. E718-E731 ◽  
Author(s):  
B. Marchetti ◽  
M. C. Morale ◽  
P. Paradis ◽  
M. Bouvier
Keyword(s):  
Hormonal Regulation ◽  
Adrenergic Receptor ◽  
Radioligand Binding ◽  
Hormonal Control ◽  
Sex Steroid ◽  
Rat Tissues ◽  
Female Rat ◽  
Adrenergic Agonists ◽  
Binding Studies ◽  
Beta 2

In the present study, we have characterized the beta 2-adrenergic receptor (beta 2-AR)-adenosine 3',5'-cyclic monophosphate (cAMP) system of the rat thymus gland and examined the hormonal regulation of the thymic beta 2-AR gene expression under physiological or pharmacological conditions accompanied by marked alterations of the sex steroid hormone milieu. We report here that membrane preparations of female rat thymic tissue contain iodocyanopindolol binding sites that exhibit pharmacological properties typical of a beta-AR. Detailed analysis by computer modeling of the binding potencies of a large series of beta 1- and beta 2-adrenergic agonists and antagonists revealed predominantly the beta 2-AR subtype (78%) in rat thymus. This inference from radioligand binding studies was corroborated functionally by the rank order of potencies of a series of adrenergic agonists to stimulate the production of cAMP. Northern blot analysis, using a human beta 2-AR cDNA as a probe, revealed the presence of a mRNA of 2.3 kb, which is consistent with the size of the beta 2-AR mRNA found in other rat tissues. Physiological regulation of specific beta 2-AR in the rat thymus was indicated by significant increases in both receptor concentration and steady-state levels of beta 2-AR mRNA during the diestrous 2 and proestrous phases of the rat estrous cycle and pregnancy, whereas castration sharply reduced beta 2-AR numbers and transcript levels within the thymus. The modulation of the thymic beta 2-AR-cAMP signaling system by the preexisting sex steroid milieu, coupled with the sex-dependent adrenergic modulation of thymic cell-mediated immune response, may contribute to the various sex-related alterations in immune responsiveness and could play a role in sexually related immune disorders.

scholarly journals Molecular determinants underlying DS2 activity at δ-containing GABAA receptors

2021 ◽  
Author(s):  
Christina B. Falk-Petersen ◽  
Frederik Rostrup ◽  
Rebekka Löffler ◽  
Stine Buchleithner ◽  
Kasper Harpsøe ◽  
...  
Keyword(s):  
Binding Site ◽  
Drug Targets ◽  
Transmembrane Domain ◽  
Direct Interaction ◽  
Binding Pocket ◽  
Docking Studies ◽  
Tonic Inhibition ◽  
Site Directed Mutagenesis ◽  
Receptor Subtypes ◽  
Molecular Determinants

AbstractDelta selective compound 2 (DS2) is one of the most widely used tools to study selective actions mediated by δ subunit-containing GABAA receptors. DS2 was discovered over 10 years ago, but despite great efforts, the precise molecular site of action has remained elusive.Using a combination of computational modeling, site-directed mutagenesis and cell-based pharmacological assays, we probed three potential binding sites for DS2 and analogs at α4β1δ receptors: an α4(+)δ(-) interface site in the extracellular domain (ECD), equivalent to the diazepam binding site in αβγ2 receptors, and two sites in the transmembrane domain (TMD); one in the α4(+)β1(-) and one in the α4(-)β1(+) interface, with the α4(-)β1(+) site corresponding to the binding site for etomidate and a recently disclosed low-affinity binding site for diazepam. We show that mutations in the ECD site did not abrogate DS2 modulation. However, mutations in the TMD α4(+)β1(-) interface, either α4(S303L) of the α4(+)-side or β1(I289Q) of the β1(-)-side, convincingly disrupted the positive allosteric modulation by DS2. This was consistently demonstrated both in an assay measuring membrane potential changes and by whole-cell patchclamp electrophysiology and rationalized by docking studies. Importantly, general sensitivity to modulators was not compromised in the mutated receptors. This study sheds important light on the long-sought molecular recognition site for DS2, refutes the misconception that the selectivity of DS2 for δ-containing receptors is caused by a direct interaction with the δ-subunit, and instead points towards a functional selectivity of DS2 and its analogs via a surprisingly well-conserved binding pocket in the TMD.Significance statementδ-Containing GABAA receptors represent potential drug targets for the treatment of several neurological conditions with aberrant tonic inhibition. Yet, no drugs are currently in clinical use. With the identification of the molecular determinants responsible for positive modulation by the know compound DS2, the ground is laid for design of ligands that selectively target δ-containing GABAA receptor subtypes, for better understanding of tonic inhibition, and, ultimately, for rational development of novel drugs.

scholarly journals Combining endocannabinoids with retigabine for enhanced M-channel effect and improved KV7 subtype selectivity

2020 ◽  
Vol 152 (8) ◽  
Author(s):  
Johan E. Larsson ◽  
Urban Karlsson ◽  
Xiongyu Wu ◽  
Sara I. Liin
Keyword(s):  
Adverse Effects ◽  
Xenopus Oocytes ◽  
Site Directed Mutagenesis ◽  
Resting Membrane Potential ◽  
Xenopus Laevis Oocytes ◽  
Kv7 Channels ◽  
Subtype Selectivity ◽  
Channel Effect ◽  
Low Concentrations ◽  
Electrode Voltage

Retigabine is unique among anticonvulsant drugs by targeting the neuronal M-channel, which is composed of KV7.2/KV7.3 and contributes to the negative neuronal resting membrane potential. Unfortunately, retigabine causes adverse effects, which limits its clinical use. Adverse effects may be reduced by developing M-channel activators with improved KV7 subtype selectivity. The aim of this study was to evaluate the prospect of endocannabinoids as M-channel activators, either in isolation or combined with retigabine. Human KV7 channels were expressed in Xenopus laevis oocytes. The effect of extracellular application of compounds with different properties was studied using two-electrode voltage clamp electrophysiology. Site-directed mutagenesis was used to construct channels with mutated residues to aid in the mechanistic understanding of these effects. We find that arachidonoyl-L-serine (ARA-S), a weak endocannabinoid, potently activates the human M-channel expressed in Xenopus oocytes. Importantly, we show that ARA-S activates the M-channel via a different mechanism and displays a different KV7 subtype selectivity compared with retigabine. We demonstrate that coapplication of ARA-S and retigabine at low concentrations retains the effect on the M-channel while limiting effects on other KV7 subtypes. Our findings suggest that improved KV7 subtype selectivity of M-channel activators can be achieved through strategically combining compounds with different subtype selectivity.

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