Prokaryotic Expression,Purification and DNA Binding Activity of DEK Protein's Carboxy-terminal DNA-binding Region
- VernacularTitle:DEK蛋白C末端DNA结合域的原核表达、纯化及其与DNA的结合活性
- Author:
Ying HUA
;
Hong-Gang HU
;
Xiang-Lei PENG
;
- Publication Type:Journal Article
- Keywords:
DEK protein Carboxy-terminal DNA binding region Prokaryotic expression Purification Electrophoretic mobility shift assay(EMSA)
- From:
China Biotechnology
2006;0(08):-
- CountryChina
- Language:Chinese
-
Abstract:
DEK protein's carboxy-terminal DNA-binding region(CBD)is a newly found DNA-binding domain of DEK,which contains several phosphorylation sites and has a close correlation with DEK protein's function in vivo and in vitro.Using prokaryotic expression system,the peptide of DEK protein's carboxy-terminal DNA-binding region(CDB)was expressed and purified.In detail,the CDB DNA fragment was constructed into pET-30a(+)vector,and E.coli BL21(DE3)competent cells were used as host cells.The fusion protein His-CBD was expressed by induction of IPTG and purified by Ni-NTA agarose.The result of SDS-PAGE showed that the molecular weight of the purified protein was about 10.7kDa.Electrophoretic mobility shift assay(EMSA)indicated that DEK-CDB prefered to bind to supercoiled form of DNA in vitro,it had similar character to the binding of whole length DEK protein with DNA.This suggested that the carboxy-terminal DNA-binding region of DEK protein might function on the binding of DEK protein to DNA partly.
