Protein Kinase C, gamma (PRKCG) (Active) Protein

Details for Product No. ABIN411969, Supplier: Log in to see
Protein Name
  • PRKCG
  • PKC-gamma
  • PKCC
  • PKCG
  • SCA14
  • PKCgamma
  • Pkcc
  • Prkcc
  • PKC
  • PKCI
  • Prkc
  • RATPKCI
  • protein kinase C gamma
  • protein kinase C, gamma
  • protein kinase C gamma type
  • PRKCG
  • Prkcg
  • LOC484316
Origin
Human
11
3
1
Source
Baculovirus infected Insect Cells
5
3
2
1
1
1
1
Protein Type
Recombinant
Biological Activity
Active
Application
Functional Studies (Func), SDS-PAGE (SDS), Western Blotting (WB)
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Purpose >90% Pure active Recombinant PKC gamma.
Characteristics Protein Source: Baculovirus (Sf9 insect cells)
Purity SDS-PAGE: ≥80 %
Background PKCγ is a member of the protein kinase C (PKC) family of serine- and threonine-specific protein kinases that can phosphorylate a wide variety of protein targets known to be involved in diverse cellular signaling pathways. In the brain, PKC is translocated to cell membranes during ischemia and is rapidly removed or degraded during the second otherwise lethal ischemic insult in preconditioned brains. This suggest that ischemic preconditioning enhances downregulation of cell signaling mediated by PKC and may thereby provide neuroprotection (1).
Alternate Names: PKC, protein kinase C, gammaprotein kinase C
Molecular Weight 105.0 kDa
Gene ID 5582
NCBI Accession NM_002739
Pathways WNT Signaling, EGFR Signaling Pathway, Neurotrophin Signaling Pathway, Thyroid Hormone Synthesis, Myometrial Relaxation and Contraction, G-protein mediated Events, Positive Regulation of Response to DNA Damage Stimulus, Interaction of EGFR with phospholipase C-gamma, Thromboxane A2 Receptor Signaling, VEGF Signaling
Application Notes Optimal working dilution should be determined by the investigator.
Comment

Activity Specification: The specific activity of PKCγ was determined to be 512 nmol /min/mg as per activity assay protocol.

Protocol Note: these are suggested working dilutions and it is recommended that the researcher perform a serial dilution of Active PKCgamma for optimal results). [ 32 P]-ATP Assay Cocktail Prepare 250μM [ 32 P]-ATP Assay Cocktail in a designated radioactive working area by adding the following components: 150 µL of 10 mM ATP Stock Solution, 100 µL [ 32P ]- ATP (0 mCi/100 µL), 5.70 ml of Kinase Assay Buffer. Store 0 ml aliquots at -20 °C. Kinase Dilution Buffer, pH 7.2 Kinase Assay Buffer I diluted at a 1:4 ratio (5X dilution) with distilled H2O. 10 mM ATP Stock Solution Prepare ATP stock solution by dissolving 50 mg of ATP in 10 ml of Kinase Assay Buffer. Store 200 µL aliquots at -20 °C. Kinase Assay Buffer I, pH 7.2 Buffer components: 20 mM MOPS, 12.0 mM beta-glycerol-phosphate, 20 mM MgC1 2, 0 mM EGTA, 0 mM EDTA. Add 0.20 mM DTT to Kinase Assay Buffer prior to use. Substrate PKC synthetic peptide substrate (ERMRPRKRQGSVRRRV) diluted in distilled H2O to a final concentration of 0 mg/mL. Assay Protocol Step 1. Thaw [ 32 P]-ATP Assay Cocktail in shielded container in a designated radioactive working area. Step 2. Thaw the Active PKCgammaamma, Kinase Assay Buffer, Substrate and Enzyme Dilution Buffer on ice. Step 3. In a pre-cooled microfuge tube, add the following reaction components bringing the initial reaction volume up to 20 µL: Component 1. 10 µL of diluted Active PKCgamma. Sonicate lipid for 1 minute prior to use.
4. Set up the blank control as outlined in step 3, excluding the addition of the substrate. Replace the substrate with an equal volume of distilled H2O.
5. Initiate the reaction by the addition of 0 µL [ 32 P]-ATP Assay Cocktail bringing the final volume up to 20 µL and incubate the mixture in a water bath at 30 °C for 15 minutes.
6. After the 15 minute incubation period, terminate the reaction by spotting 20 µL of the reaction mixture onto individual pre-cut strips of phosphocellulose P81 paper.
7. Air dry the pre-cut P81 strip and sequentially wash in a 1 % phosphoric acid solution (dilute 10 ml of phosphoric acid and make a 1L solution with distilled H2O) with constant gentle stirring. It is recommended that the strips be washed a total of 3 intervals for approximately 10 minutes each.
8. Count the radioactivity on the P81 paper in the presence of scintillation fluid in a scintillation counter.
9. Determine the corrected cpm by removing the blank control value (see Step 4) for each sample and calculate the kinase specific activity as outlined below.
Restrictions For Research Use only
Format Liquid
Concentration 0.1 mg/mL
Buffer Recombinant protein stored in 50 mM Tris-HCl, pH 7.5, 150 mM NaCl, 0.25 mM DTT, 0.1 mM EGTA, 0.1 mM EDTA, 0.1 mM PMSF, 25 % glycerol.
Preservative Dithiothreitol (DTT)
Precaution of Use This product contains dithiothreitol (DTT): a POISONOUS AND HAZARDOUS SUBSTANCE which should be handled by trained staff only.
Handling Advice Centrifuge the vial prior to opening.
Storage -80 °C
Storage Comment -80°C
Expiry Date 12 months
Supplier Images
Western Blotting (WB) image for Protein Kinase C, gamma (PRKCG) (Active) protein (ABIN411969) Protein Kinase C, gamma (PRKCG) (Active) protein
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