ChemiSCREEN™ Human α_2A Adrenergic Receptor Membrane Preparation

ChemiSCREEN™ Human α_2A Adrenergic Receptor Membrane Preparation

• ChemiScreen
• Chemicon (Millipore)

200 units

Full-length human ADRA2A transcript variant 1 cDNA encoding α_2A

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_2A 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_2A. The membrane preparations exhibit a Kd of 5.4 nM for [³H]-MK-912. With 5 nM [³H]-MK-912, 5 µg/well α_2A Membrane Prep typically yields greater than 20-fold signal-to-background ratio.

Radioligand binding assay and GTPγS binding.

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. Studies in mouse revealed that both the alpha2A and alpha2C subtypes were required for normal presynaptic control of transmitter release from sympathetic nerves in the heart and from central noradrenergic neurons; the alpha2A subtype inhibited transmitter release at high stimulation frequencies, whereas the alpha2C subtype modulated neurotransmission at lower levels of nerve activity. This gene encodes alpha2A subtype and it contains no introns in either its coding or untranslated sequences.

FUNCTION: SwissProt: P08913 # 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 oxymetazoline > clonidine > epinephrine > norepinephrine > phenylephrine > dopamine > p-synephrine > p-tyramine > serotonin = p-octopamine. For antagonists, the rank order is yohimbine > phentolamine = mianserine > chlorpromazine = spiperone = prazosin > propanolol > alprenolol = pindolol.
SIZE: 450 amino acids; 48957 Da
SUBCELLULAR LOCATION: Cell membrane; Multi-pass membrane protein.
SIMILARITY: SwissProt: P08913 ## Belongs to the G-protein coupled receptor 1 family.

Human

Chemicon

alpha2A

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 MK-912 at 5 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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Adrenergic Receptors are G-Protein Coupled Receptors that stimulate cell growth and proliferation. Mutant isoforms of the β-2 Adrenergic Receptor are associated with asthma, obesity and diabetes.

• ADRA2A
• ZNF32
• ADRA2R
• ALPHA2AAR
• Alpha-2AAR
• ADRA2
• OTTHUMP00000058849
• ADRAR

GPCR Membrane Preparations

ChemiSCREEN™ Human α_2A Adrenergic Receptor Membrane Preparation

alpha2A

Table 1. Signal:background and specific binding values obtained in a competition binding assay with α_2A membrane prep and unlabeled rauwolscine.

	10 µg/well	5 µg/well
Signal:Background	28.97	34.14
Specific Binding (cpm)	11777	9147

SPECIFICATIONS: 1 unit = 5 µg
Bmax for [3H] MK-912 binding: 71.4 pmol/mg protein
Kd for [3H] MK-912 binding: ~5.4 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] MK-912 (Perkin Elmer#: NET-1059)
Wash Buffer: 50 mM Hepes, pH 7.4, 500mM NaCl , 0.1% BSA, filtered and stored at 4°C.

Chem-1

P08913

NM_000681.2

GPCR