Purification and activity assay of HSA-AX15 (R13K) fusion protein expressed in Pichia pastoris.
- Author:
Hong-Liang ZHAO
1
;
Chong XUE
;
Xiang-Hua XIONG
;
Wei ZHANG
;
Bing-Fen YANG
;
Zhi-Min LIU
Author Information
1. Beijing Institute of Biotechnology, Beijing 100071, China.
- Publication Type:Journal Article
- MeSH:
Animals;
Ciliary Neurotrophic Factor;
genetics;
Humans;
Mice;
Mutant Proteins;
biosynthesis;
genetics;
Pichia;
genetics;
metabolism;
Recombinant Fusion Proteins;
biosynthesis;
genetics;
isolation & purification;
Serum Albumin;
genetics
- From:
Chinese Journal of Biotechnology
2005;21(2):254-258
- CountryChina
- Language:Chinese
-
Abstract:
To increase the in vivo half-life of human CNTF mutein AX15 (R13K), HSA-AX15 (R13K) fusion protein was constructed by the fusion of the C-terminus of HSA to the N-terminus of AX15 (R13K) via an 11 amino acids linker. HSA-AX15 (R13K) fusion protein was purified to homogeneity by cation exchange chromatography, reverse phase chromatography and gel filtration after expressed in pichia pastoris. TF-1 cell survival bioassay showed the biological activity of AX15 (R13K) was not affected by the fusion to HSA. It was demonstrated that tertian injection of 4.8 mg/kg HSA-AX15 (R13K) fusion protein could produce more potent anti-obesity effects on KM mice than daily injection of 1.6 mg/kg AX15 (R13K). The long-acting form of hCNTF variant has the potential to reduce discomfort by requiring fewer injections and to minimize the side-effects by decreasing the dosage and fluctuation of plasma concentration, and thus has superior clinical application.
