Secretory Expression of the Fusion Protein PTH-HSA in Pichia pastoris
- VernacularTitle:人甲状旁腺激素-血清白蛋白融合蛋白的构建及在毕赤酵母中的表达
- Author:
Jun WANG
;
Wei SHEN
;
Zhi-Ming RAO
;
Ge-Jian ZHU
;
- Publication Type:Journal Article
- Keywords:
Human parathyroid hormone Fusion protein Vector pPIC9K Pichia pastoris
- From:
China Biotechnology
2006;0(02):-
- CountryChina
- Language:Chinese
-
Abstract:
The fused gene (PTH-HSA) of parathyroid hormone (PTH) gene and Human Serum Albumin(HSA) gene was amplified without linker by Overlapping PCR technology. The spliced gene was clone into Pichia pastoris secretory vector pPIC9K. With the help of promoter AOX1 and mat ? signal peptide, the PTH-HSA gene was designed to secretory expression.Linearized by restriction enzyme SalI, The recombinant plasmid pPIC9K/PTH-HSA was transformed into Pichia pastoris KM71 by electroporation. The recombinant strains which were identified by G418 and PCR analysis were induced by methanol to express protein PTH-HSA. The target protein was expressed in fermentation supernatant. Western blot analysis of the fusion protein showed that the expressed fusion protein PTH-HSA had the antigenicity of HSA.adenylate cyclase assay proved that the fused protein exhibited the bioactivity to stimulate cAMP synthesis The specific activity of broth was about 318IU/ml.
