Biological Information

Background Information:

Glycogen synthase kinase-3 (GSK-3) is a protein kinase that was originally identified as a regulator of glycogen synthase, a key enzyme in glycogen metabolism. There are two isoforms of GSK-3, GSK-3a and GSK-3b, and they are strictly regulated via phosphorylation. Phosphorylation of GSK-3b on Ser9 by protein kinase B (PKB) causes its inactivation and is the primary mechanism responsible for growth factor inhibition of this kinase. Activation of GSK-3b is dependent upon the phosphorylation of Tyr216. Upon activation, it has been shown to phosphorylate a number of different cellular proteins, including p53, c-Myc, c-Jun, heat shock factor-1 (HSF-1), and cyclin D1. GSK-3b also has been shown to phosphorylate aberrant sites on the microtubule associated protein tau, which is critical for the progression of Alzheimer’s disease. The inhibitors may have therapeutic opportunities for the treatment of diabetes, Alzheimer's disease, stroke and bipolar disorder.

Family:

Kinase

Sub Family:

CMGC

Class:

Protein Ser/Thr

Protein Name:

GSK3beta

Uniprot Number:

P49841

Protein Aliases:

Serine/threonine-protein kinase GSK3B

Gene Name:

GSK3B

Gene ID:

2932

Accession Number:

NM_001146156

Organism:

Human

Construct Details:

Fragment (2-end), Recombinant

Assay Information

Assay Type:

Biochemical

Assay Sub Type:

Enzymatic

Functional Mode:

Antagonist

Detection Method:

Radiometry

Measured Response:

Scintillation

Testing Information

Procedure Summary:

Human recombinant protein kinase GSK-3b expressed in Sf21 insect cells is used. Test compound and/or vehicle is preincubated with 500 ng/ml enzyme‡ in modified MOPS buffer pH 7.2 for 15 minutes at 37⁰C. The reaction is initiated by addition of 5 μM phospho-glycogen synthase peptide-2, 10 μM ATP and 0.25 μCi [γ32P]ATP for another 30 minute incubation period and terminated by further addition of 3% H3PO4. An aliquot is removed and counted to determine the amount of [32P]Phospho-glycogen synthase peptide-2 formed. Compounds are screened at 10 μM.

Substrate:

5 µM Phospho-glycogen synthase peptide-2

ATP Concentration:

10 uM

Incubation:

30 min at 37°C

Control Inhibitor:

5-iodo-Indirubin-3'-monoxime

Criteria For Significance:

≥ 50% of max stimulation or inhibition

Test Concentration / Dose:

Suggested concentration for initial testing is 10 uM

Test Sample Requirements:

Minimum for 1) Screen: 25 μl of 10 mM stock -OR- 1 mg (pre-weighed). 2) Dose Response: 60 μl of 10 mM stock -OR- 1 mg (pre-weighed).

Minimum Order Quantity:

2

Turnaround Time

Standard:

15 Business Days

Reference Compound Data

PubChem ID

176158

Name

SB 216763

Measurement

0.0027μM - IC50

PubChem ID

4210951

Name

SB 415286

Measurement

0.065μM - IC50

PubChem ID

5472313

Name

5-iodo-Indirubin-3'-monoxime

Measurement

0.0018μM - IC50

PubChem ID

6539732

Name

GSK-3 Inhibitor II

Measurement

0.15μM - IC50

PubChem ID

122130844

Name

Staurosporine

Measurement

0.006μM - IC50

Clinical Relevance

Therapeutic Area:

Oncology/Immuno-Oncology

Additional Information

Brand:

LeadHunter

Testing Location:

Taiwan - Taipei