title: [Expression and bioactivity of OSF-1 in Pichia pastoris].
abstract: In order to research the biologic activity of osteoblast-stimulating factor 1 (OSF-1), the pPIC9K/osf-1 yeast expression vector was constructed to express and purify OSF-1. Firstly, the osf-1 gene sequence was obtained by artificial synthesis and cloned into Pichia pastoris expression vector pPIC9K to generate pPIC9K/osf-1. The recombinant plasmid was linearized by Sac I and transformed into P. pastoris GS115 by electroporation. Recombinant P. pastoris GS115/ pPIC9K/osf-1 was screened by MD and G418-YPD plates and further identified by PCR. The positive P. pastoris was induced with 1% methanol at 25 degrees C for 96 h. The target protein was analyzed by SDS-PAGE showing a special band about 18 kDa. The target protein was successfully purified from the supernatant of the broth using ion exchange chromatography of SP-Sephadex C-50. The purity of target protein was above 98%. Western blotting appeared a good antigenicity of the purified protein. Bioassay results show that the recombinant protein OSF-1 can promote the differentiation and proliferation of osteoblasts MC3T3-E1. We successfully expressed OSF-1 by recombinant P. pastoris for further development of anti-osteoporosis of research and industrial production of OSF-1.
authors.forename: Donghong
authors.initials: D
authors.lastname: Kang
authors.forename: Yan
authors.initials: Y
authors.lastname: Wang
authors.forename: Hongmei
authors.initials: H
authors.lastname: Zhang
authors.forename: Xiaoyu
authors.initials: X
authors.lastname: Feng
authors.forename: Wei
authors.initials: W
authors.lastname: Cao
authors.forename: Ping
authors.initials: P
authors.lastname: Wang
mesh_terms: D015153:Blotting, Western; D002352:Carrier Proteins / Q000096:biosynthesis*; D002846:Chromatography, Affinity; D016207:Cytokines / Q000096:biosynthesis*; D004591:Electrophoresis, Polyacrylamide Gel; D018274:Electroporation; D010843:Pichia / Q000378:metabolism*; D016133:Polymerase Chain Reaction; D011994:Recombinant Proteins / Q000096:biosynthesis
chemical_list: D002352:Carrier Proteins; D016207:Cytokines; D011994:Recombinant Proteins; C066845:pleiotrophin