| Purification and Quality Control | The recombinant protein is purified by affinity chromatography in combination with FPLC columns. The purified Positive Cofactor 4 (PC4) F77P mutation is greater than 90% homogeneous based on SDS-PAGE analysis. |
| Unit Definition (Activity) | 1 unit equals 1 nanogram of purified protein. 1 unit is sufficient for a gel mobility shift assay in a 20 µl reaction; 100 units are sufficient for protein-protein interaction assays. |
| Applications | Recombinant PC4 has been utilized for in vitro function studies, including transcription, DNA replication, in vitro phosphorylation, gel mobility shift assay, protein-protein interactions and as a substrate for in vitro acetylation. |
| Formulation and Storage | The protein is in 20mM Tris-HCl pH7.9,100mM NaCl, 0.2mM EDTA, 1mM DTT and 20% glycerol. Stored at -70°C before use. Avoid repeated freeze thaw cycles. |
| Synonym | Homo sapiens SUB1 homolog (S. cerevisiae) (SUB1); MGC102747; p14 and P15. |
| Protein Sequence | MPKSKELVSS GSSGSDSDSE VDKKLKRKKQ VAPEKPVKKQ KTGETSRALS SSKQSSSSRD
DNMFQIGKMR YVSVRDPKGK VLIDIREYWM DPEGEMKPGR KGISLNPEQW SQLKEQISDI
DDAVRKL |
| Background | The human PC4 is a non-TAF transcription coactivator that mediates activator-dependent transcription by RNA polymerase II in vitro through most tested activators. The function of PC4 is through interactions with transcriptional activators and the basal transcription machinery. It is negatively regulated by casein kinase II phosphorylation both in vitro and in vivo (1-3). PC4 strongly binds single stranded DNA and the region essential for the single stranded DNA binding activity was mapped around residue 77. A single amino acid change at position 77 (F to P) abolishes both ds- and ss-DNA binding activity (4-6). |
| References | 1. Ge, H. et al., (1994) Cell 78, 513-523; (1994) Proc. Natl. Acad. Sci. USA 91, 12691-12695.
2. Kretzschmar, M. et al., (1994) Cell 78, 525-534
3. Wu, SY. et al., (1998) J. Biol. Chem. 273, 12492-12498
4. Brandsen, J. et al., (1997) Nat. Struct. Biol. 4, 900-903
5. Werten, S. et al., (1998) EMBO J. 17, 5103-5111
6. Ge, H. (2000) Nucleic Acids Res. 28, e3 |
