title: [Expression of fusion protein of parathyroid hormone and transferrin N-terminal half-molecule in Pichia pastoris].
abstract: The fused gene (PTH-TFN) of parathyroid hormone (PTH) gene and transferring N-terminal half-molecule (TFN) gene was amplified by multiple PCR and inserted into pPIC9 vector. The recombinant plasmid pPIC9-PTH-TFN was transformed into Pichia pastoris GS115 by PEG. After methanol induction, the target protein was expressed in fermentation supernatant at high level. The fused protein PTH-TFN with purity being higher than 95% was finally obtained after purification through two-step chromatography: SP Sepharose Fast Flow and Phenyl Sepharose Fast Flow. Western blot analysis and adenylate cyclase assay proved that the fused protein exhibited the bioactivity to stimulate cAMP synthesis and the ability to bind Fe3+ in the Fe3+ saturation study as the recombinant TFN did indicating that TFN could be used as the transcellar carrier of PTH.
authors.affiliation: Institute of Biotechnology, Academy of Military Medical Sciences, Beijing 100071, China.
authors.forename: Hao
authors.initials: H
authors.lastname: Zhang
authors.forename: Xiao-Jing
authors.initials: XJ
authors.lastname: Li
authors.forename: De-Jie
authors.initials: DJ
authors.lastname: Wang
authors.forename: Jing
authors.initials: J
authors.lastname: Chen
authors.forename: Yan-Ying
authors.initials: YY
authors.lastname: Li
authors.forename: Yu-Ling
authors.initials: YL
authors.lastname: Li
authors.forename: Ming-Shan
authors.initials: MS
authors.lastname: Shen
authors.forename: Hong-Qing
authors.initials: HQ
authors.lastname: Fang
authors.forename: Hui-Peng
authors.initials: HP
authors.lastname: Chen
mesh_terms: D019677:Artificial Gene Fusion; D003001:Cloning, Molecular; D006801:Humans; D010281:Parathyroid Hormone / Q000235:genetics*; D010843:Pichia / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis* / Q000235:genetics; D014168:Transferrin / Q000235:genetics*
chemical_list: C031138:N-terminal monoferrictransferrin; C470125:PTH protein, human; D010281:Parathyroid Hormone; D011993:Recombinant Fusion Proteins; D014168:Transferrin