title: High-level expression of a manganese superoxide dismutase (PoMn-SOD) from Pleurotus ostreatus in Pichia pastoris.
abstract: The full-length cDNA of Pleurotus ostreatus superoxide dismutase (PoMn-SOD) was cloned and successfully expressed by using the pPIC9K vector under the control of alcohol oxidase 1 promoter with a secretion signal peptide (α-factor) in Pichia pastoris. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and Western blotting demonstrated that recombinant PoMn-SOD, a 21.8 kDa protein, was secreted into the culture medium. Nondenaturing PAGE experiments confirmed that recombinant PoMn-SOD was secreted in a functionally active form and the expression system did not require any acid activation process. The factors affecting the expression level were optimized in shaking flask cultures. The maximum enzyme activity (156.9 U/mg) was observed under the following conditions: Initial medium pH was 6.0, induction time point was at the 6th day, and methanol concentration was 0.7 % (v/v). This was the first report on secretory expression of recombinant PoMn-SOD in P. pastoris, which might provide a reference for further practical applications.
authors: Key Laboratory of Agro-Microbial Resources and Utilization, Ministry of Agriculture, College of Food Science and Technology, Huazhong Agricultural University, Wuhan, 430070, China.
authors: Chaomin
authors: C
authors: Yin
authors: Wenxia
authors: W
authors: Zhao
authors: Liesheng
authors: L
authors: Zheng
authors: Liguo
authors: L
authors: Chen
authors: Qi
authors: Q
authors: Tan
authors: Xiaodong
authors: X
authors: Shang
authors: Aimin
authors: A
authors: Ma
mesh_terms: D005656:Fungal Proteins / Q000096:biosynthesis* / Q000235:genetics; D015870:Gene Expression; D010843:Pichia / Q000235:genetics / Q000378:metabolism*; D020076:Pleurotus / Q000201:enzymology / Q000235:genetics*; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics; D013482:Superoxide Dismutase / Q000096:biosynthesis* / Q000235:genetics
chemical_list: D005656:Fungal Proteins; D011994:Recombinant Proteins; D013482:Superoxide Dismutase