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[Expression and purification of FOXO1 DNA binding domain and its DNA properties]
Yinuer Ha1, Jun Li1, Yongheng Chen1
1Key Laboratory of Cancer Proteomics of Chinese Ministry of Health, Xiangya Hospital, Central South University, Changsha 410008, China.
Objective:
To express and purify FOXO1 DNA binding domain (DBD) and to evaluate its DNA binding properties. Methods: Gene sequence of FOXO1-DBD was optimized and the recombinant strains were induced at low temperature to obtain soluble protein of FOXO1-DBD. Then, FOXO1-DBD protein was purified through Ni affinity chromatography and cation exchange. Finally, the DNA binding properties of FOXO1-DBD was evaluated by electrophoretic mobility shift assay (EMSA). Results: Most of soluble proteins were obtained by optimizing its genes sequence and induction at 21 ℃. Over 95% purity of FOXO1-DBD protein was obtained through two-steps purification. Purified protein exhibited good binding property to the DNA binding motif (G/ATAAACA) of FOX family. Conclusion: An efficient expression and purification method for FOXO1-DBD is established, and the complexity of FOXO1 protein in recognition of DNA is verified.
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