Human GRK6 Baculovirus-Insect cells Overexpression Lysate

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Human GRK6 Baculovirus-Insect cells Overexpression Lysate: Product Information

Product Description
This Human GRK6 overexpression lysate was created in Baculovirus-Insect cells and intented for use as a Western blot (WB) positive control. Purification of GRK6 protein (Cat: 11752-H20B) from the overexpression lysate was verified.
Expression Host
Baculovirus-Insect cells
Species
Human
Sequence Information
A DNA sequence encoding the human GRK6 isofrom B (P43250-2) (Met 1-Arg 589) was fused with the N-terminal polyhistidine-tagged GST tag at the N-terminus.
Molecule Mass
The recombinant human GRK6/GST chimera consists of 826 amino acids and has a calculated molecular mass of 95.1 kDa. It migrates as an approximately 85 kDa band in SDS-PAGE under reducing conditions.

Human GRK6 Baculovirus-Insect cells Overexpression Lysate: Usage Guide

Preparation Method
Cell lysate was prepared by homogenization of the over-expressed cells in ice-cold modified RIPA Lysis Buffer with cocktail of protease inhibitors (Sigma). Cell debris was removed by centrifugation. Protein concentration was determined by Bradford assay (Bio-Rad protein assay, Microplate Standard assay). The cell lysate was boiled for 5 min in 1 x SDS loading buffer (50 mM Tris-HCl pH 6.8, 12.5% glycerol, 1% sodium dodecylsulfate, 0.01% bromophenol blue) containing 5% b-mercaptoethanol, and lyophilized.
Lysis Buffer
Modified RIPA Lysis Buffer: 50 mM Tris-HCl pH 7.4, 150 mM NaCl, 1mM EDTA, 1% Triton X-100, 0.1% SDS, 1% Sodium deoxycholate, 1mM PMSF.
Recommend Usage
1.  Centrifuge the tube for a few seconds and ensure the pellet at the bottom of the tube. 2.  Re-dissolve the pellet using 200μL pure water and boil for 2-5 min.
Sample Buffer
1 X Sample Buffer (1 X modified RIPA buffer+1 X SDS loading buffer).
Stability & Storage
Store at 4℃ for up to twelve months from date of receipt. After re-dissolution, aliquot and store at -80℃ for up to twelve months. Avoid repeated freeze-thaw cycles.
Application
Western Blot (WB)
Optimal dilutions/concentrations should be determined by the end user.

Human GRK6 Baculovirus-Insect cells Overexpression Lysate: Alternative Names

Human GPRK6 Overexpression Lysate

GRK6 Background Information

G protein-coupled receptor kinase 6, also known as G protein-coupled receptor kinase GRK6, GRK6 and GPRK6, is a lipid-anchor protein which belongs to theprotein kinase superfamily, AGC Ser/Thr protein kinase family and GPRK subfamily. GRK6 / GPRK6 contains oneAGC-kinase C-terminal domain, oneprotein kinase domain and oneRGS domain. This protein phosphorylates the activated forms of G protein-coupled receptors thus initiating their deactivation. Several transcript variants encoding different isoforms have been described. GRK6 / GPRK6 is widely expressed. GRK6 / GPRK6 silencing causes suppression of signal transducer and activator of transcription 3 (STAT3) phosphorylation associated with reduction in MCL1 levels and phosphorylation, illustrating a potent mechanism for the cytotoxicity of GRK6 inhibition in multiple myeloma (MM) tumor cells. GRK6 also appears to be involved in responses to morphine. Inhibition of GRK6 represents a uniquely targeted novel therapeutic strategy in human multiple myeloma.
Full Name
G protein-coupled receptor kinase 6
References
  • Ghadessy,R.S. et al., 2003, Br J Pharmacol  138 (4):660-70.
  • Murga,C. et al., 2009, Brain Behav Immun. 23 (1):16-7.
  • Tiedemann,R.E. et al., 2010, Blood  115 (8):1594-604.
  • Ahmed,M.R. et al., 2010, Sci Transl Med 2 (28): 28ra28.
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