| Description | Tumor protein p53, a nuclear protein, plays an essential role in the regulation of cell cycle, specifically in the transition from G0 to G1. It is found in very low levels in normal cells, however, in a variety of transformed cell lines, it is expressed in high amounts, and believed to contribute to transformation and malignancy. p53 is a DNA-binding protein containing DNA-binding, oligomerization and transcription activation domains. It is postulated to bind as a tetramer to a p53-binding site and activate expression of downstream genes that inhibit growth and/or invasion, and thus function as a tumor suppressor.(1-5) Mutants of p53 that frequently occur in a number of different human cancers fail to bind the consensus DNA binding site, hence cause the loss of tumor suppressor activityis and highly correlated with tumorigenesis.(6, 7) The wild type His tagged p53 (393 amino acids) was expressed in a baculovirus system and purified by affinity and FPLC chromatography. Purified protein is greater than 95% homogeneous based on SDS-PAGE gel analysis. | | Target Gene Sequence | EAPRMPEAAP PVAPAPAAPT PAAPAPAPSW PLSSSVPSQK TYQGSYGFRL GFLHSGTAKS
VTCTYSPALN KMFCQLAKTC PVQLWVDSTP PPGTRVRAMA IYKQSQHMTE VVRRCPHHER
CSDSDGLAPP QHLIRVEGNL RVEYLDDRNT FRHSVVVPYE PPEVGSDCTT IHYNYMCNSS
CMGGMNRRPI LTIITLEDSS GNLLGRNSFE VRVCACPGRD RRTEEENLRK KGEPHHELPP
GSTKRALPNN TSSSPQPKKK PLDGEYFTLQ IRGRERFEMF RELNEALELK DAQAGKEPGG
SRAHSSHLKS KKGQSTSRHK KLMFKTEGPD SD | | Storage and Formulation | Protein is stored in 20 mM Tris-Cl (pH 8.0), 20% Glycerol, 100 mM KCl, 1 mM DTT and 0.2 mM EDTA buffer and kept at -80°C. Avoid repeatly freeze thaw cycles. | | Application | Recombinant p53 can be used for: 1) gel mobility shift assay or for a DNase I footprinting assay in the presence of double stranded DNA containing a consensus p53-binding sequence [5’-PuPuPuC(A/T)(T/A)GPyPyPy-3’]; 2) in vitro transcription assay; 3) protein-protein interaction assay; and 4) for cell growth assay; 5) P53 is an excellent substrate for kinase assays. 1 unit equals 1 nanogram of purified protein. 1 unit is sufficient for a gel mobility shift assay in a 20 µl reaction; 50 units are sufficient for reconstituted transcription assay and 100 units are sufficient for a protein-protein interaction assay. For research use only. | | Reference | 1. Finlay, C. et al., (1989) Cell 57, 1083-1093
2. Michalovitz, D. et al., (1990) Cell 62, 671-680
3. Baker, S. et al., (1990) Science 249, 912-915
4. Fields, S. et.al., (1990) Science 249, 1046-1049
5. Raycroft, L. et al., (1990) Science 249, 1049-1051
6. Hollstein, M. et al., (1991) Science 253, 49-53
7. Bennett, W. et al., (1992) Chest 101, 19S-20S
8. Xu et al., (2005) Nat Cell Bio, 7:165-171 |
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