ChemiScreen™ Membrane Prep Human Recombinant mGIu6

Species	Host Cells	Protein Target
Human	Chem-1	mGlu6

Species

Host Cells

Protein Target

Human

Chem-1

mGlu6

Description
Catalogue Number	HTS149M
Trade Name	ChemiScreen Chemicon
Description	ChemiSCREEN™ Membrane Preparation Recombinant Human mGlu₆ Metabotropic Glutamate Receptor
Overview	human GRM6 cDNA
Background Information	Glutamate is a main excitatory neurotransmitter in the central nervous system, and it plays a role in learning, memory and neurotoxicity. The biological actions of glutamate are mediated by ionotropic and metabotropic glutamate receptors, which are ion channels and GPCRs respectively. Metabotropic glutamate receptors (mGluRs) are members of the class 3 G-protein coupled receptor family, which are characterized by a large extracellular domain. They are further classified into group I, II, and III mGluRs on the basis of their sequence identity, pharmacology, and signal transduction mechanism. Group I (mGlu₁ and mGlu₅) couple to the phospholipase C pathway through G_αq, whereas group II (mGlu₂ and mGlu₃) and group III (mGlu₄, mGlu₆, mGlu₇, and mGlu₈) negatively couple to the adenylyl cyclase pathway though G_αi (Conn and Pin, 1997). mGlu₆ has a restricted localization to the dendrites of ON bipolar cells that synapse with retinal rods and cones. Targeted deletion of mGlu₆ results in deficits in ON responses but unchanged OFF responses to light (Masu et al., 1995). In addition, mutations in GRM6 in humans are responsible for some forms of autosomal recessive congenital stationary night blindness (Dryja et al., 2005; Zeitz et al., 2005). Chemicon's mGlu₆ 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 mGlu₆ interactions with its ligands. The cell line exhibits a calcium response with EC50s of 6.2 µM, 23.4 µM, and 52.7 µM for L-AP4, (RS)PPG, and L-glutamate. The membrane preparations exhibit EC50s of of 4.0 µM, 24.1 µM, and 49.3 µM for L-AP4, (RS)PPG, and L-glutamate, respectively, in a GTPγS binding assay.

Description

Catalogue Number

HTS149M

Trade Name

ChemiScreen
Chemicon

Description

ChemiSCREEN™ Membrane Preparation Recombinant Human mGlu₆ Metabotropic Glutamate Receptor

Overview

human GRM6 cDNA

Background Information

Glutamate is a main excitatory neurotransmitter in the central nervous system, and it plays a role in learning, memory and neurotoxicity. The biological actions of glutamate are mediated by ionotropic and metabotropic glutamate receptors, which are ion channels and GPCRs respectively. Metabotropic glutamate receptors (mGluRs) are members of the class 3 G-protein coupled receptor family, which are characterized by a large extracellular domain. They are further classified into group I, II, and III mGluRs on the basis of their sequence identity, pharmacology, and signal transduction mechanism. Group I (mGlu₁ and mGlu₅) couple to the phospholipase C pathway through G_αq, whereas group II (mGlu₂ and mGlu₃) and group III (mGlu₄, mGlu₆, mGlu₇, and mGlu₈) negatively couple to the adenylyl cyclase pathway though G_αi (Conn and Pin, 1997). mGlu₆ has a restricted localization to the dendrites of ON bipolar cells that synapse with retinal rods and cones. Targeted deletion of mGlu₆ results in deficits in ON responses but unchanged OFF responses to light (Masu et al., 1995). In addition, mutations in GRM6 in humans are responsible for some forms of autosomal recessive congenital stationary night blindness (Dryja et al., 2005; Zeitz et al., 2005). Chemicon's mGlu₆ 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 mGlu₆ interactions with its ligands. The cell line exhibits a calcium response with EC50s of 6.2 µM, 23.4 µM, and 52.7 µM for L-AP4, (RS)PPG, and L-glutamate. The membrane preparations exhibit EC50s of of 4.0 µM, 24.1 µM, and 49.3 µM for L-AP4, (RS)PPG, and L-glutamate, respectively, 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 with no preservatives. Packaging method: Membrane protein was adjusted to 1 mg/ml in 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 with no preservatives.
Packaging method: Membrane protein was adjusted to 1 mg/ml in packaging buffer, rapidly frozen, and stored at -80°C.

Applications
Application	Human mGlu6 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 mGlu6 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_000843.3
Entrez Gene Summary	L-glutamate is the major excitatory neurotransmitter in the central nervous system and activates both ionotropic and metabotropic glutamate receptors. Glutamatergic neurotransmission is involved in most aspects of normal brain function and can be perturbed in many neuropathologic conditions. The metabotropic glutamate receptors are a family of G protein-coupled receptors, that have been divided into 3 groups on the basis of sequence homology, putative signal transduction mechanisms, and pharmacologic properties. Group I includes GRM1 and GRM5 and these receptors have been shown to activate phospholipase C. Group II includes GRM2 and GRM3 while Group III includes GRM4, GRM6, GRM7 and GRM8. Group II and III receptors are linked to the inhibition of the cyclic AMP cascade but differ in their agonist selectivities.
Gene Symbol	GRM6 DKFZp686H1993 GPRC1F mGluR6 MGLUR6 CSNB1B mGlu6
GPCR Class	C
Host Cells	Chem-1
Protein Target	mGlu6
Target Sub-Family	Glutamate (metabotropic)
UniProt Number	O15303
UniProt Summary	FUNCTION: SwissProt: O15303 # Receptor for glutamate. The activity of this receptor is mediated by a G-protein that inhibits adenylate cyclase activity. SIZE: 877 amino acids; 95437 Da SUBCELLULAR LOCATION: Cell membrane; Multi-pass membrane protein. DISEASE: SwissProt: O15303 # Defects in GRM6 are the cause of congenital stationary night blindness type 1B (CSNB1B) [MIM:257270]. This disorder consits of a previously unrecognized, autosomal recessive form of congenital night blindness associated with a negative electroretinogram waveform. Patients are night blind from an early age, and when maximally dark-adapted, they could perceive lights only with an intensity equal to or slightly dimmer than that normally detected by the cone system. ERGs in response to single brief flashes of light have clearly detectable a-waves, which are derived from photoreceptors, and greatly reduced b-waves, which are derived from the second-order inner retinal neurons. ERGs in response to sawtooth flickering light indicate a markedly reduced ON response and a nearly normal OFF response. There is no subjective delay in the perception of suddenly appearing white vs black objects on a gray background. SIMILARITY: SwissProt: O15303 ## Belongs to the G-protein coupled receptor 3 family.
Incubation 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 DCG IV, (2R4R) APDC, and glutamate are 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 L-AP4, (RS)PPG, or glutamate -stimulated [³⁵S]-GTPγS binding. The mGlu₆ 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_000843.3

Entrez Gene Summary

L-glutamate is the major excitatory neurotransmitter in the central nervous system and activates both ionotropic and metabotropic glutamate receptors. Glutamatergic neurotransmission is involved in most aspects of normal brain function and can be perturbed in many neuropathologic conditions. The metabotropic glutamate receptors are a family of G protein-coupled receptors, that have been divided into 3 groups on the basis of sequence homology, putative signal transduction mechanisms, and pharmacologic properties. Group I includes GRM1 and GRM5 and these receptors have been shown to activate phospholipase C. Group II includes GRM2 and GRM3 while Group III includes GRM4, GRM6, GRM7 and GRM8. Group II and III receptors are linked to the inhibition of the cyclic AMP cascade but differ in their agonist selectivities.

Gene Symbol

GRM6
DKFZp686H1993
GPRC1F
mGluR6
MGLUR6
CSNB1B
mGlu6

GPCR Class

Host Cells

Chem-1

Protein Target

mGlu6

Target Sub-Family

Glutamate (metabotropic)

UniProt Number

O15303

UniProt Summary

FUNCTION: SwissProt: O15303 # Receptor for glutamate. The activity of this receptor is mediated by a G-protein that inhibits adenylate cyclase activity.
SIZE: 877 amino acids; 95437 Da
SUBCELLULAR LOCATION: Cell membrane; Multi-pass membrane protein.
DISEASE: SwissProt: O15303 # Defects in GRM6 are the cause of congenital stationary night blindness type 1B (CSNB1B) [MIM:257270]. This disorder consits of a previously unrecognized, autosomal recessive form of congenital night blindness associated with a negative electroretinogram waveform. Patients are night blind from an early age, and when maximally dark-adapted, they could perceive lights only with an intensity equal to or slightly dimmer than that normally detected by the cone system. ERGs in response to single brief flashes of light have clearly detectable a-waves, which are derived from photoreceptors, and greatly reduced b-waves, which are derived from the second-order inner retinal neurons. ERGs in response to sawtooth flickering light indicate a markedly reduced ON response and a nearly normal OFF response. There is no subjective delay in the perception of suddenly appearing white vs black objects on a gray background.
SIMILARITY: SwissProt: O15303 ## Belongs to the G-protein coupled receptor 3 family.

Incubation 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 DCG IV, (2R4R) APDC, and glutamate are 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 L-AP4, (RS)PPG, or glutamate -stimulated [³⁵S]-GTPγS binding.
The mGlu₆ 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	EC50 in GTPγS binding assay by Glutamate: ~ 49.3 μM EC50 in GTPγS binding assay by (RS)PPG: ~ 24.1 μM EC50 in GTPγS binding assay by L-AP4: ~ 4.04 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

EC50 in GTPγS binding assay by Glutamate: ~ 49.3 μM
EC50 in GTPγS binding assay by (RS)PPG: ~ 24.1 μM
EC50 in GTPγS binding assay by L-AP4: ~ 4.04 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

办公用品

ChemiScreen™ Membrane Prep Human Recombinant mGIu6

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