| Description | Human p53 protein is composed of 393 amino acid residues with several distinct regions. The N-terminal activation domain allows p53 protein to recruit the basal transcription machinery and activate the expression of target genes. The core domain binds to target DNA in a sequence-specific manner and the majority of mutations found in human tumors occur in the region of the gene encoding this domain (1-3). The C-terminal domain is composed of predominantly basic residues and modification of the C-terminal basic domain, including acetylation, glycosylation and phosphorylation, is an essential mechanism for regulating p53 function (4-6). Disruption or loss of oligomerization function is associated with loss of cell cycle arrest (5, 6). This mutant protein (with the deletion of the C-terminus 51 residues including the entire basic domain and a portion of the tetramerization domain) can be used as a unique tool to study specific functions of p53 related to the C-terminus.
The His tagged C-terminus-deleted p53 (amino acid 1-342) was expressed in a baculovirus system and purified by affinity and FPLC chromatography.
Recombinant p53 can be used for: 1) gel mobility shift assay or for a DNase I footprinting 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) cell growth assay. | | Gene Sequence | EPQSDPSVEP PLSQETFSDL WKLLPENNVL SPLPSQAMDD LMLSPDDIEQ WFTEDPGPDE
APRMPEAAPP VAPAPAAPTP AAPAPAPSWP LSSSVPSQKT YQGSYGFRLG FLHSGTAKSV
TCTYSPALNK MFCQLAKTCP VQLWVDSTPP PGTRVRAMAI YKQSQHMTEV VRRCPHHERC
SDSDGLAPPQ HLIRVEGNLR VEYLDDRNTF RHSVVVPYEP PEVGSDCTTI HYNYMCNSSC
MGGMNRRPIL TIITLEDSSG NLLGRNSFEV RVCACPGRDR RTEEENLRKK GEPHHELPPG
STKRALPNNT SSSPQPKKKP LDGEYFTLQI RGRERFEMF | | Formulation and Storage | This protein was kept in 20 mM Tris-Cl (pH 8.0), 20% glycerol, 100 mM KCl, 1 mM DTT and 0.2 mM EDTA. Always store at -80˚C, avoid repeated freeze thaw cycles. | | Application | 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. Variable in different lots. For Research Use Only. | | References | 1. Pellegata, NS. et al., (1995) Oncogene 11, 337-349
2. El-Deiry, WS. et al., (1992) Nature Genet 1, 45-49
3. Hollstein, M. et al., (1991) Science 253, 49-53
4. Hupp, TR. et al., (1992) Cell 71, 875-886
5. Ishioka, C. et al., (1995) Oncogene 10, 1485-1492
6. Waterman, MJ. et al., (1996) Cancer Res 56, 158-163 |
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