| Purification and Quality Control | The His-tag recombinant protein is purified by affinity chromatography in combination with FPLC columns. The purified SV40 T-Antigen is greater than 95% homogeneous based on SDS-PAGE analysis. | | Unit Definition (Activity) | 1 unit equals 1 nanogram of purified protein. 50-500 units are sufficient for an in vitro unwinding assay and 100 units are sufficient for a protein-protein interaction assay. | | Applications | Recombinant SV40 T antigen can be used for: 1) DNA unwinding assay; 2) DNA replication assay; 3) DNA binding assays; 4) protein-protein interaction assays; and 5) cell growth assay. | | 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. | | Protein Sequence | MDKVLNREES LQLMDLLGLE RSAWGNIPLM RKAYLKKCKE FHPDKGGDEE KMKKMNTLYK
KMEDGVKYAH QPDFGGFWDA TEIPTYGTDE WEQWWNAFNE ENLFCSEEMP SSDDEATADS
QHSTPPKKKR KVEDPKDFPS ELLSFLSHAV FSNRTLACFA IYTTKEKAAL LYKKIMEKYS
VTFISRHNSY NHNILFFLTP HRHRVSAINN YAQKLCTFSF LICKGVNKEY LMYSALTRDP
FSVIEESLPG GLKEHDFNPE EAEETKQVSW KLVTEYAMET KCDDVLLLLG MYLEFQYSFE
MCLKCIKKEQ PSHYKYHEKH YANAAIFADS KNQKTICQQA VDTVLAKKRV DSLQLTREQM
LTNRFNDLLD RMDIMFGSTG SADIEEWMAG VAWLHCLLPK MDSVVYDFLK CMVYNIPKKR
YWLFKGPIDS GKTTLAAALL ELCGGKALNV NLPLDRLNFE LGVAIDQFLV VFEDVKGTGG
ESRDLPSGQG INNLDNLRDY LDGSVKVNLE KKHLNKRTQI FPPGIVTMNE YSVPKTLQAR
FVKQIDFRPK DYLKHCLERS EFLLEKRIIQ SGIALLLMLI WYRPVAEFAQ SIQSRIVEWK
ERLDKEFSLS VYQKMKFNVA MGIGVLDWLR NSDDDDEDSQ ENADKNEDGG EKNMEDSGHE
TGIDSQSQGS FQAPQSSQSV HDHNQPYHIC RGFTCFKKPP TPPPEPET | | Background | The large T antigen (TAg) of Simian virus 40 acts as an initiator of DNA replication, participates in cellular transformation, and induces cell growth (1-4). A number of functional domains have been characterized including the N-terminal J domain, nuclear localization signal (NLS), internal helicase domain, Rb and topoisomerase I interacting domain, p53 and ATP binding domain (5,6). The p53-interacting domain is also essential for binding p300/CBP (7). Phosphorylation of TAg by several kinases may regulate its function both positively and negatively (8). | | References | 1. Fanning E., (1992) J. Viol. 66, 1289-1293.
2. Pan ZQ. et al., (1996) J. Biol. Chem. 271, 22111-22116.
3. Chao, HA. et al., (2000) Mol. Cell. Biol. 20, 7624-7633.
4. Quartin, RS. et al., (1994) J. Viol. 68, 1334-1341.
5. Marton, A. et al., (1993) Nucleic Acids Res. 21, 1689-1695.
6. Kim, HY. et al., (2001) EMBO J. 20, 295-304.
7. Lill, NL. et al., (1997) Nature 387, 823-827.
8. Moarefi, IF. et al., (1993) J. Viol. 67, 4992-5002. |
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