Rb-C Fusion Protein #6022
Inquiry Info. # 6022
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Product Specifications
| MW (kDa) | 76 |
Product Information
Product Description
Rb-C (retinoblastoma protein C-terminus) Fusion Protein serves as a useful substrate for various cyclin-dependent kinases (CDK's) (8,9,11). It is expressed as a recombinant protein fusion of Rb residues 701-928 and maltose binding protein. The phosphorylation sites present in this C-terminal portion of Rb include Ser780 (8), Ser795 (9) and Ser807/811 (11).
| MW (kDa) | 76 |
| Molecular Formula | Apparent Molecular Weight: 68 kDa |
Product Usage Information
Rb-C Fusion Protein at a concentration of 2 µg/20 µl reaction can be phosphorylated using cdc2 Protein Kinase (NEB #P6020) in an in vitro kinase assay with 1X Kinase Buffer #9802 and 200 µM ATP #9804. After 30 minutes at 30ºC, phosphorylation can be detected by Western blot with Phospho-Rb (Ser795) Antibody #9301.
Quality Control
The purified protein was resolved on two identical SDS-polyacrylamide gels. One was stained with Coomassie brilliant blue and the other was blotted to PVDF membrane and the protein band detected using Rb antibody. Greater than 95% of the observable protein was identified as the Rb-C Fusion Protein by apparent molecular weight (68 kDa), and only one band was identified by immunoblotting.
Source / Purification
Recombinant protein fusion of Rb residues 701-928 and maltose binding protein. Purified by the pMAL Protein Purification System (New England Biolabs #E8000S).
Background
The retinoblastoma tumor suppressor protein Rb regulates cell proliferation by controlling progression through the restriction point within the G1-phase of the cell cycle (1). Rb has three functionally distinct binding domains and interacts with critical regulatory proteins including the E2F family of transcription factors, c-Abl tyrosine kinase, and proteins with a conserved LXCXE motif (2-4). Cell cycle-dependent phosphorylation by a CDK inhibits Rb target binding and allows cell cycle progression (5). Rb inactivation and subsequent cell cycle progression likely requires an initial phosphorylation by cyclin D-CDK4/6 followed by cyclin E-CDK2 phosphorylation (6). Specificity of different CDK/cyclin complexes has been observed in vitro (6-8) and cyclin D1 is required for Ser780 phosphorylation in vivo (9).
- Sherr, C.J. (1996) Science 274, 1672-7.
- Nevins, J.R. (1992) Science 258, 424-9.
- Welch, P.J. and Wang, J.Y. (1993) Cell 75, 779-90.
- Hu, Q.J. et al. (1990) EMBO J 9, 1147-55.
- Knudsen, E.S. and Wang, J.Y. (1997) Mol Cell Biol 17, 5771-83.
- Lundberg, A.S. and Weinberg, R.A. (1998) Mol Cell Biol 18, 753-61.
- Connell-Crowley, L. et al. (1997) Mol Biol Cell 8, 287-301.
- Kitagawa, M. et al. (1996) EMBO J 15, 7060-9.
- Geng, Y. et al. (2001) Proc Natl Acad Sci USA 98, 194-9.
Alternate Names
p105-Rb; pp110; retinoblastoma; screeningkit; substrate
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For Research Use Only. Not for Use in Diagnostic Procedures.
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