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Human CAMK1G / CLICK III / CaMKIgamma Insect Cell Lysate (WB positive control)

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Human CAMK1G/CLICK III Transfected / Overexpression Cell Lysate Product Information
Expressed Host:Baculovirus-Insect cells
Product Description:Baculovirus-Insect Cell lysate that Human CAMK1G / CLICK III / CaMKIgamma transfected / overexpressed for Western blot (WB) positive control. The whole cell lysate is provided in 1X Sample Buffer (1X modified RIPA buffer+1X SDS loading buffer).
Sequence information:A DNA sequence encoding the human CAMK1G isoform 1 (Q96NX5-1) (Met 1-Met 476) was fused with the N-terminal polyhistidine-tagged GST tag at the N-terminus.
Predicted N Terminal:Met
Molecule Mass:The recombinant human CAMK1G/GST chimera consists of 713 amino acids and has a calculated molecular mass of 81 kDa. It migrates as an approximately 75 kDa band in SDS-PAGE under reducing conditions.
Human CAMK1G/CLICK III Transfected / Overexpression Cell Lysate Usage Guide
Preparation Method:Cell lysate was prepared by homogenization 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.
Quality Control Testing:12.5% SDS-PAGE Stained with Coomassie Blue after protein purification.
Stability:Samples are stable for up to twelve months from date of receipt.
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. 3. Store the lyophilized cell lysate at 4℃. After re-dissolution, recommend to aliquot it into smaller quantities and store at -80℃.
Storage Buffer:1 X Sample Buffer (1 X modified RIPA buffer+1 X SDS loading buffer).
Storage Instruction:Store at 4℃. After re-dissolution, aliquot and store at -80℃.
Application notes:Western blot (WB): Use at an assay dependent dilution.
Other Applications: Not tested.
Optimal dilutions/concentrations should be determined by the end user.
CAMK1G/CLICK III/CaMKIgamma Background

Calmodulin-Dependent Protein Kinase (CaM Kinase) is a kind of protein phosphorylate multiple downstream targets. Concentration of cytosolic calcium functions as a second messenger that mediates a wide range of cellular responses. Calcium binds to calcium binding proteins (calmodulin/CaM) and stimulates the activity of a variety of enzymes, including CaM kinases referred to as CaM-kinases (CaMKs), such as CaMKI, CaMKII, CaMKIV and CaMKK. Calmodulin-dependent protein kinase CL3/CaMKIγ is a memberane-anchored CaMK belonging to the CaM kinase family. Its C-terminal region is uniquely modified by two sequential lipidification steps: prenylation followed by a kinase-activity-regulated palmitoylation. These modifications are essential for CaMKIγ membrane anchoring and targeting into detergent-resistant lipid microdomains in the dendrites. It has been found that CaMKIγ critically contributed to BDNF-stimulated dendritic growth. Raft insertion of CaMKIγ specifically promoted dendritogenesis of cortical neurons by acting upstream of RacGEF STEF and Rac, both present in lipid rafts. Thus, CaMKIγ may represent a key element in the Ca2+-dependent and lipid-raft-delineated switch that turns on extrinsic activity-regulated dendrite formation in developing cortical neurons. 

Human CAMK1G/CLICK III/CaMKIgamma References
  • Davare MA, et al. (2009) Transient receptor potential canonical 5 channels activate Ca2+ / calmodulin kinase Igamma to promote axon formation in hippocampal neurons. J Neurosci. 29 (31): 9794-808.
  • Takemoto-Kimura S, et al. (2007) Regulation of dendritogenesis via a lipid-raft-associated Ca2+ / calmodulin - dependent protein kinase CLICK-III / CaMKIgamma. Neuron. 54 (5): 755-70.
  • Harrill JA, et al. (2010) Splice variant specific increase in Ca2+ / calmodulin-dependent protein kinase 1-gamma mRNA expression in response to acute pyrethroid exposure. J Biochem Mol Toxicol. 24 (3): 174-86.
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    Catalog: 13148-H20BL-300
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