Biochemical characterization of manganese superoxide dismutase from Deinococcus radiodurans

• Main Authors: Yien-Ting Tsai, 蔡彥廷
• Other Authors: Chih-Cheng Lin, Min Yuan Chou
• Format: Others
• Language: zh-TW
• Published: 2008
• Online Access: http://ndltd.ncl.edu.tw/handle/rk283e

碩士 === 元培科技大學 === 生物技術研究所 === 96 === Abstract The enzyme manganese superoxide dismutase(Mn-SOD) is one of the antioxidant enzymes involved in cellular defense against oxidative stress and catalyzes the conversion of O.2- into the stabler H2O2. In this study, a putative gene encoding Mn-SOD from Denococcus radiodurans(dMn-SOD) was cloned, sequensed, expressed in Escherichia coli Bl21 (DE3) and the protein was purified using HiTrap column. Sequencing resulted ORF of 635bp, which corresponded to 211 amino acid. Recombinant protein was dissolved in SDS-PAGE and it was analyzed that its weight was 23.4kD. The fusion protein had 50 U/mg activity. It was active in a range of basic pH(from 7.0 to 9.0). When EDTA was added in protein solution, its activity was down. In addation, using the site-direct mutagenesis changed the amino acid of active center, its activity was going down. Result of study was suggested that Deinococcus radiodurans(DEIRA) soda gene was cloned in Escherichia coli BL21 to produce Mn-SOD what receives the pH value, chelating agent, and influence of sour change of the amine base near active in the center.