Product Pathways - Tyrosine Kinase/ Adaptors
HER2/ErbB2 Kinase #7382
Cell Signaling Technology offers a full line of protein kinases, substrates, antibody detection reagents and HTScan® kits. Browse our "Reagents for High-Throughput Screening" product listing or contact us at drugdiscovery@cellsignal.com.
Description
Purified recombinant human HER2/ErbB2 (Lys676-Val1255) kinase, supplied as a GST fusion protein.
Source / Purification
The GST-Kinase fusion protein was produced using a baculovirus expression system with a construct expressing human HER2/ErbB2 (Lys676-Val1255) (GenBank Accession No. NM_004448) with an amino-terminal GST tag. The protein was purified by one-step affinity chromatography using glutathione-agarose.
Gel Staining
Figure 1. The purity of the GST-HER2/ErbB2 fusion protein was analyzed using SDS/PAGE followed by Coomassie stain.
Kinase Assay - Radiometric
Figure 2. HER2/ErbB2 kinase activity was measured in a radiometric assay using the following reaction conditions: 5 mM MOPS, pH 7.2, 2.5 mM β-glycerophosphate, 5 mM MnCl2, 1 mM EGTA, 0.4 mM EDTA, 4 mM MgCl2, 0.05 mM DTT, 50 μM ATP, Substrate: Poly(Glu-Tyr), 400 ng/μL, and recombinant HER2/ErbB2: variable.
Quality Control
The theoretical molecular weight of the GST-HER2/ErbB2 fusion protein is 116 kDa. The purified kinase was quality controlled for purity using SDS-PAGE followed by Coomassie stain [Fig.1]. HER2/ErbB2 kinase activity was determined using a radiometric assay [Fig.2].
Background
The ErbB2 (HER2) proto-oncogene encodes a 185 kDa transmembrane, receptor-like glycoprotein with intrinsic tyrosine kinase activity (1). While ErbB2 lacks an identified ligand, ErbB2 kinase activity can be activated in the absence of a ligand when overexpressed and through heteromeric associations with other ErbB family members (2). Amplification of the ErbB2 gene and overexpression of its product are detected in almost 40% of human breast cancers (3). Binding of the c-Cbl ubiquitin ligase to ErbB2 at Tyr1112 leads to ErbB2 poly-ubiquitination and enhances degradation of this kinase (4). ErbB2 is a key therapeutic target in the treatment of breast cancer and other carcinomas with the regulation of ErbB2 degradation by the c-Cbl-regulated proteolytic pathway as one potential therapeutic strategy.Phosphorylation of the kinase domain residue Tyr877 of ErbB2 (homologous to Tyr416 of pp60c-Src) may be involved in regulating ErbB2 biological activity. The major autophosphorylation sites in ErbB2 areTyr1248 and Tyr1221/1222; phosphorylation of these sites couples ErbB2 to the Ras-Raf-MAP kinase signal transduction pathway (1,5).
- Muthuswamy, S.K. et al. (1999) Mol. Cell. Biol. 19, 6845-6857.
- Qian, X. et al. (1994) Proc. Natl. Acad. Sci. USA 91, 1500-1504.
- Dittadi, R. and Gion, M. (2000) J. Natl. Cancer Inst. 92, 1443-1444.
- Klapper, L.N. et al. (2000) Cancer Res. 60, 3384-3388.
- Kwon, Y.K. et al. (1997) J. Neurosci. 17, 8293-8299.
Application References
Have you published research involving the use of our products? If so we'd love to hear about it. Please let us know!
Companion Products
- 7058 HTScan® HER2/ErbB2 Kinase Assay Kit
- 1305 FLT3 (Tyr589) Biotinylated Peptide
- 9411 Phospho-Tyrosine Mouse mAb (P-Tyr-100)
- 9805 HTScan® Tyrosine Kinase Buffer (4X)
- 9804 ATP (10 mM)
- 9953 Staurosporine
- 7450 Tyrosine Kinase Substrate Screening Kit
This product is for in vitro research use only and is not intended for use in humans or animals. This product is not intended for use as therapeutic or in diagnostic procedures.
