ChemiScreen™ Recombinant Human Glucagon Receptor Membrane Preparation

ChemiScreen™ Recombinant Human Glucagon Receptor Membrane Preparation

• ChemiScreen
• Chemicon (Millipore)

200 units

TRANSFECTION: Full-length human GCGR cDNA encoding Glucagon Receptor
HOST CELLS: Chem-1, an adherent mammalian cell line with no endogenous glucagon receptor expression.

Glucagon is a 29-amino acid peptide that stimulates glycogenolysis and gluconeogenesis in the liver to increase blood glucose. The receptor for glucagon is a class 2 (or class B) GPCR that signals through G_s to stimulate cAMP production (Mayo et al., 2003). Mice lacking the glucagon receptor have mild hypoglycemia after fasting, and exhibit hyperplasia of pancreatic α-cells (Gelling et al., 2003). Because of its role in promoting hyperglycemia, the glucagon receptor presents a potential target for treatment of diabetes. Chemicon's glucagon receptor 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 antagonists of glucagon receptor interactions and its ligands. The membrane preparations exhibit a Kd of 3.09 nM for [¹²⁵I]-glucagon. With 1 nM [¹²⁵I]-glucagon, 5 or 10 μg/well glucagon receptor Membrane Prep yields greater than 10-fold signal-to-background ratio.

Radioligand binding assay and GTPγS binding

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.

The physiologic effects of glucagon (GCG; MIM 138030) are mediated through the glucagon receptor, a member of the superfamily of receptors characterized by a 7-transmembrane domain structure and by their coupling via GTP-binding proteins (G proteins) to adenyl cyclase.[supplied by OMIM]

FUNCTION: SwissProt: P47871 # This is a receptor for glucagon which plays a central role in regulating the level of blood glucose by controlling the rate of hepatic glucose production and insulin secretion. The activity of this receptor is mediated by G proteins which activate adenylyl cyclase and also a phosphatidylinositol-calcium second messenger system.
SIZE: 477 amino acids; 54009 Da
SUBCELLULAR LOCATION: Cell membrane; Multi-pass membrane protein.
DISEASE:SwissProt: P47871 # It is a candidate for certain defects in non-insulin- dependent diabetes mellitus (NIDDM). The variant in position 40 (Ser) is found in some NIDDM patients, but also in nondiabetic subjects.
SIMILARITY: SwissProt: P47871 ## Belongs to the G-protein coupled receptor 2 family.

Human

Chemicon

glucagon / GCG

Liquid in packaging buffer: 50 mM Tris pH 7.4, 10% glycerol and 1% BSA; no preservatives.
Packaging method: Membrane proteins were adjusted to 0.5 mg/ml in 1 ml packaging buffer, rapidly frozen, and stored at -80°C.

peptide

Glucagon

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

B

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• GCGR
• MGC138246
• GL-R
• GGR

GPCR Membrane Preparations

ChemiScreen™ Recombinant Human Glucagon Receptor Membrane Preparation

SPECIFICATIONS:
1 unit = 5 μg
B_max for [¹²⁵I] glucagon binding: 16.7 pmol/mg protein
K_d for [¹²⁵I] glucagon binding: ~ 3.09 nM

Signal:background and specific binding values obtained in a competition binding assay with varying amounts of Glucagon receptor membrane prep:

	10 µg/well	5 µg/well
Signal:Background	10.1	35
Specific Binding (cpm)	33315	36252

Chem-1

RECOMMENDED ASSAY CONDITIONS: Membranes are mixed with radioactive ligand and unlabeled competitor in binding buffer in a nonbinding 96-well plate, and incubated for 1-2 h. Prior to filtration, a GF/C 96-well filter plate 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: [¹²⁵I]-glucagon (Perkin Elmer#:NEX-207)
• Wash Buffer: 50 mM Hepes, pH 7.4, 500mM NaCl , 0.1% BSA, filtered and stored at 4°C.

One package contains enough membranes for at least 200 assays (units), where a unit is the amount of membrane that will yield greater than 10-fold signal:background with [¹²⁵I]-labeled glucagon at 1 nM

Chem-1

P47871

NM_000160.2

GPCR