ChemiScreen™ Membrane Prep Hu Recombinant GPR109A

Species	Host Cells	Protein Target
Human	Chem-4	GPR109A

Species

Host Cells

Protein Target

Human

Chem-4

GPR109A

Description
Catalogue Number	HTS201M
Trade Name	ChemiScreen Chemicon
Description	ChemiSCREEN™ Membrane Preparation Recombinant Human GPR109A Receptor
Overview	Full-length human GPR109A cDNA
Background Information	Nicotinic acid (niacin), a vitamin of the B complex, is used clinically in high doses to decrease total plasma levels of cholesterol. Interestingly, the total cholesterol levels and low-density lipoprotein (LDL) concentrations decrease, while the high-density lipoprotein (HDL) concentrations increase with nicotinic acid treatment. The cholesterol-lowering effects of nicotinic acid result from inhibition of lipolysis in adipose tissue, which decreases plasma levels of free fatty acid (FFA) (Altschul et al., 1955; Carlson, 1963). In a study of nicotinic acid and myocardial infarction in the Coronary Drug Project (1966-1975), patients receiving 3 g/day nicotinic acid exhibited reduced rates of myocardial infarction (Coronary Drug Project Research Group, 1975). However, an unwanted effect of high doses of nicotinic acid is vasodilatation, occurring mainly in the upper body and face, known as flushing. Recently two receptors for nicotinic acid have been identified as class A G protein-coupled receptors that couple to Gi to inhibit accumulation of cAMP (Offermans, 2006). GPR109A (also known as HM74A in humans and PUMA-G in mice) is a high affinity receptor for nicotinic acid, whereas GPR109B (also known as HM74) is a low affinity receptor for nicotinic acid that is found in humans but not rodents (Wise et al., 2003). GPCR109A is found primarily in adipose tissue and immune cells. Millipore's GPR109A 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 GPR109A interactions with its ligands. The membrane preparations exhibit EC50s of 3.65 μM for nicotinic acid in a GTPγS binding assay.

Description

Catalogue Number

HTS201M

Trade Name

ChemiScreen
Chemicon

Description

ChemiSCREEN™ Membrane Preparation Recombinant Human GPR109A Receptor

Overview

Full-length human GPR109A cDNA

Background Information

Nicotinic acid (niacin), a vitamin of the B complex, is used clinically in high doses to decrease total plasma levels of cholesterol. Interestingly, the total cholesterol levels and low-density lipoprotein (LDL) concentrations decrease, while the high-density lipoprotein (HDL) concentrations increase with nicotinic acid treatment. The cholesterol-lowering effects of nicotinic acid result from inhibition of lipolysis in adipose tissue, which decreases plasma levels of free fatty acid (FFA) (Altschul et al., 1955; Carlson, 1963). In a study of nicotinic acid and myocardial infarction in the Coronary Drug Project (1966-1975), patients receiving 3 g/day nicotinic acid exhibited reduced rates of myocardial infarction (Coronary Drug Project Research Group, 1975). However, an unwanted effect of high doses of nicotinic acid is vasodilatation, occurring mainly in the upper body and face, known as flushing. Recently two receptors for nicotinic acid have been identified as class A G protein-coupled receptors that couple to Gi to inhibit accumulation of cAMP (Offermans, 2006). GPR109A (also known as HM74A in humans and PUMA-G in mice) is a high affinity receptor for nicotinic acid, whereas GPR109B (also known as HM74) is a low affinity receptor for nicotinic acid that is found in humans but not rodents (Wise et al., 2003). GPCR109A is found primarily in adipose tissue and immune cells. Millipore's GPR109A 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 GPR109A interactions with its ligands. The membrane preparations exhibit EC50s of 3.65 μM for nicotinic acid in a GTPγS binding assay.

Product Information
Format	Membranes
Presentation	Liquid in packaging buffer: 50 mM Tris pH 7.4, 10% glycerol and 1% BSA no preservatives. Packaging method: Membranes protein were adjusted to 1 mg/ml in 1 ml packaging buffer, rapidly frozen, and stored at -80°C

Product Information

Format

Membranes

Presentation

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

Applications
Application	Human GPR109A GPCR membrane preparation for Radioligand binding Assays & GTPγS binding.
Key Applications	Radioligand binding assay GTPγS binding
Application Notes	Radioligand binding assay and GTPγS binding.

Applications

Application

Human GPR109A GPCR membrane preparation for Radioligand binding Assays & GTPγS binding.

Key Applications

Radioligand binding assay
GTPγS binding

Application Notes

Radioligand binding assay and GTPγS binding.

Biological Information
Entrez Gene Number	NM_177551.3
Gene Symbol	GPR109A HM74a HM74A PUMAG Puma-g HM74b
GPCR Class	A
Host Cells	Chem-4
Protein Target	GPR109A
Target Sub-Family	Nicotinic Acid
UniProt Number	Q8TDS4
UniProt Summary	FUNCTION: SwissProt: Q8TDS4 # Acts as a high affinity receptor for nicotinic acid and mediates its anti-lipolytic effect through a G(i)-protein-mediated inhibition of adenylyl cyclase. This pharmacological effect requires nicotinic acid doses that are much higher than those provided by a normal diet. SIZE: 363 amino acids; 41850 Da SUBCELLULAR LOCATION: Cell membrane; Multi-pass membrane protein. TISSUE SPECIFICITY: Expression largely restricted to adipose tissue and spleen. SIMILARITY: SwissProt: Q8TDS4 ## Belongs to the G-protein coupled receptor 1 family. MISCELLANEOUS: The rank order of potency for the displacement of nicotinic acid binding is 5-methyl pyrazole-3-carboxylic acid = pyridine-3-acetic acid > acifran > 5-methyl nicotinic acid = acipimox > nicotinuric acid = nicotinamide.
Incubation Conditions	ASSAY CONDITIONS: Membranes are permeabilized by addition of saponin to an equal concentration by mass, then mixed with [³⁵S]-GTPγS (final concentration of 0.3 nM) in 20 mM HEPES, pH 7.4/100 mM NaCl/10 mM MgCl2/0.5 μM GDP in a nonbinding 96-well plate. Unlabeled nicotinic acid was added to the final concentration indicated in Figure 1 (final volume 100 μL), and incubated for 30 min at 30°C. The binding reaction is transferred to a GF/B filter plate (Millipore MAHF B1H) previously prewetted with water, and washed 3 times (1 mL per well per wash) with cold 10 mM sodium phosphate, pH 7.4. The plate is dried and counted. One vial contains enough membranes for at least 200 assays (units), where one unit is the amount of membrane that will yield greater than 1000 cpm specific nicotinic acid-stimulated [³⁵S]-GTPγS binding. The GPR109A membrane preparation is expected to be functional in a radioligand binding assay; however, the end user will need to determine the optimal radiolabeled ligand for use with this product.

Biological Information

Entrez Gene Number

NM_177551.3

Gene Symbol

GPR109A
HM74a
HM74A
PUMAG
Puma-g
HM74b

GPCR Class

Host Cells

Chem-4

Protein Target

GPR109A

Target Sub-Family

Nicotinic Acid

UniProt Number

Q8TDS4

UniProt Summary

FUNCTION: SwissProt: Q8TDS4 # Acts as a high affinity receptor for nicotinic acid and mediates its anti-lipolytic effect through a G(i)-protein-mediated inhibition of adenylyl cyclase. This pharmacological effect requires nicotinic acid doses that are much higher than those provided by a normal diet.
SIZE: 363 amino acids; 41850 Da
SUBCELLULAR LOCATION: Cell membrane; Multi-pass membrane protein.
TISSUE SPECIFICITY: Expression largely restricted to adipose tissue and spleen.
SIMILARITY: SwissProt: Q8TDS4 ## Belongs to the G-protein coupled receptor 1 family.
MISCELLANEOUS: The rank order of potency for the displacement of nicotinic acid binding is 5-methyl pyrazole-3-carboxylic acid = pyridine-3-acetic acid > acifran > 5-methyl nicotinic acid = acipimox > nicotinuric acid = nicotinamide.

Incubation Conditions

ASSAY CONDITIONS: Membranes are permeabilized by addition of saponin to an equal concentration by mass, then mixed with [³⁵S]-GTPγS (final concentration of 0.3 nM) in 20 mM HEPES, pH 7.4/100 mM NaCl/10 mM MgCl2/0.5 μM GDP in a nonbinding 96-well plate. Unlabeled nicotinic acid was added to the final concentration indicated in Figure 1 (final volume 100 μL), and incubated for 30 min at 30°C. The binding reaction is transferred to a GF/B filter plate (Millipore MAHF B1H) previously prewetted with water, and washed 3 times (1 mL per well per wash) with cold 10 mM sodium phosphate, pH 7.4. The plate is dried and counted.

One vial contains enough membranes for at least 200 assays (units), where one unit is the amount of membrane that will yield greater than 1000 cpm specific nicotinic acid-stimulated [³⁵S]-GTPγS binding.

The GPR109A membrane preparation is expected to be functional in a radioligand binding assay; however, the end user will need to determine the optimal radiolabeled ligand for use with this product.

Product Usage Statements
Quality Assurance	SPECIFICATIONS: 1 unit = 5 μg EC50 in GTPγS binding assay by Nicotinic Acid: ~ 3.65 μM
Usage Statement	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.

Product Usage Statements

Quality Assurance

SPECIFICATIONS: 1 unit = 5 μg
EC50 in GTPγS binding assay by Nicotinic Acid: ~ 3.65 μM

Usage Statement

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.

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

Storage and Shipping Information

Storage Conditions

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

Packaging Information
Material Size	200 units

Packaging Information

Material Size

200 units

实验耗材

分子生物学试剂

蛋白与免疫学

生化试剂

细胞及检测试剂

色谱及层析

仪器设备

实验服务

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ChemiScreen™ Membrane Prep Hu Recombinant GPR109A

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