ChemiSCREEN™ Human α_2B Adrenergic Receptor Membrane Preparation

ChemiSCREEN™ Human α_2B Adrenergic Receptor Membrane Preparation

• ChemiScreen
• Chemicon (Millipore)

200 units

Full-length human ADRA2B transcript cDNA encoding α_2B.

The endogenous catecholamines epinephrine and norepinephrine have profound effects on smooth muscle activity, cardiac function, carbohydrate and fat metabolism, hormone secretion, neurotransmitter release, and central nervous system actions. These activities are mediated by GPCRs belonging to two subfamilies, the α- and β-adrenoceptors (Bylund et al., 1994). The α₂ adrenergic receptor subfamily members, consisting of α_2A, α_2B, and α_2C, couple primarily to Gi to inhibit cAMP production, and play an important role in regulation of cardiovascular and CNS function. Experiments with α_2A-selective agonists and mice lacking α_2A demonstrate that activation of α_2A results in hypotension, sedation, analgesia, and hypothermia (Kable et al., 2000). Chemicon's α_2B membrane preparations are crude membrane preparations made from our proprietary stable recombinant cell lines to ensure high-level of GPCR surface expression; thus, they are ideal HTS tools for screening of agonists and antagonists at α_2B. The membrane preparations exhibit a Kd of 11.5 nM for [³H]-Rauwolscine. With 15 nM [³H]-Rauwolscine, 5 µg/well α_2B Membrane Prep typically yields greater than 30-fold signal-to-background ratio.

Human

Unless otherwise stated in our catalog or other company documentation accompanying the product(s), our products are intended for research use only and are not to be used for any other purpose, which includes but is not limited to, unauthorized commercial uses, in vitro diagnostic uses, ex vivo or in vivo therapeutic uses or any type of consumption or application to humans or animals.

Alpha-2-adrenergic receptors are members of the G protein-coupled receptor superfamily. They include 3 highly homologous subtypes: alpha2A, alpha2B, and alpha2C. These receptors have a critical role in regulating neurotransmitter release from sympathetic nerves and from adrenergic neurons in the central nervous system. This gene encodes the alpha2B subtype, which was observed to associate with eIF-2B, a guanine nucleotide exchange protein that functions in regulation of translation. A polymorphic variant of the alpha2B subtype, which lacks 3 glutamic acids from a glutamic acid repeat element, was identified to have decreased G protein-coupled receptor kinase-mediated phosphorylation and desensitization; this polymorphic form is also associated with reduced basal metabolic rate in obese subjects and may therefore contribute to the pathogenesis of obesity. This gene contains no introns in either its coding or untranslated sequences.

FUNCTION: SwissProt: P18089 # Alpha-2 adrenergic receptors mediate the catecholamine- induced inhibition of adenylate cyclase through the action of G proteins. The rank order of potency for agonists of this receptor is clonidine > norepinephrine > epinephrine = oxymetazoline > dopamine > p-tyramine = phenylephrine > serotonin > p-synephrine / p-octopamine. For antagonists, the rank order is yohimbine > chlorpromazine > phentolamine > mianserine > spiperone > prazosin > alprenolol > propanolol > pindolol.
SIZE: 450 amino acids; 49954 Da
SUBCELLULAR LOCATION: Cell membrane; Multi-pass membrane protein.
SIMILARITY: SwissProt: P18089 ## Belongs to the G-protein coupled receptor 1 family.

Human

Chemicon

alpha2B

One package contains enough membranes for at least 200 assays (units), where a unit is the amount of membrane that will yield greater than 20-fold signal:background with 3H-labeled Rauwolscine at 15 nM

Liquid in packaging buffer: 50 mM Tris pH 7.4, 10% glycerol and 1% BSA with no preservatives.

non-peptide

Maintain frozen at -70°C for up to 2 years. Do not freeze and thaw.

Adrenergic

A

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• ADRA2B
• ADRA2L1
• ALPHA2BAR
• ADRA2RL1
• ADRARL1

GPCR Membrane Preparations

ChemiSCREEN™ Human α_2B Adrenergic Receptor Membrane Preparation

alpha2B

Table 1. Signal:background and specific binding values obtained in a competition binding assay with α_2B membrane prep and unlabeled rauwolscine.

	5 µg/well
Signal:Background	64.7
Specific Binding (cpm)	14646

SPECIFICATIONS: 1 unit = 5 µg
Bmax for [3H]-rauwolscine binding: 154 pmol/mg protein
Kd for [3H]-rauwolscine binding: ~11.5 nM

Chem-1

Membranes are mixed with radioactive ligand and unlabeled competitor (see Figures 1 and 2 for concentrations tested) in binding buffer in a nonbinding 96-well plate, and incubated for 1-2 h. Prior to filtration, an FC 96-well harvest plate (Millipore cat. # MAHF C1H) is coated with 0.33% polyethyleneimine for 30 min, then washed with 50mM HEPES, pH 7.4, 0.5% BSA. Binding reaction is transferred to the filter plate, and washed 3 times (1 mL per well per wash) with Wash Buffer. The plate is dried and counted.

Binding buffer: 50 mM Hepes, pH 7.4, 5 mM MgCl2, 1 mM CaCl2, 0.2% BSA, filtered and stored at 4°C
Radioligand: [3H]-Rauwolscine (Perkin Elmer#: NET-722)
Wash Buffer: 50 mM Hepes, pH 7.4, 500mM NaCl , 0.1% BSA, filtered and stored at 4°C.

Chem-1

P18089

NM_000682.5

GPCR