为了阐明过氧化物酶EfeB对大肠杆菌胞内氧化应激调控的影响,对编码基因efeB进行了同源重组敲除及同源过表达,考察了菌体生长、丙二醛浓度及胞内活性氧水平的变化,并进一步比较了efeB过表达菌株中与氧化胁迫相关基因efeU、efeO、sodA、oxyR、katE和细菌重组修复基因recA在正常和过氧化氢刺激下的表达水平变化。结果表明:过表达efeB对菌体生长有促进作用,过表达菌株Eco/pEE胞内的丙二醛含量相比出发菌株(223.79 nmol/g蛋白)下降了85%,胞内活性氧是出发菌株的1.36倍,同时katE下调了87.62%,efeU、efeO、sodA、oxyR、recA分别上调3.88、1.33、2.48、1.95和1.49倍,过表达菌株Eco/pEE在过氧化氢刺激条件下,katE进一步下调了94.81%,recA进一步上调了8.09倍;反之,菌株缺失efeB生长减缓,胞内丙二醛的含量是出发菌株的1.15倍,胞内活性氧是出发菌株的0.5倍。以上结果提出了大肠杆菌EfeB在细胞氧化应激下新的生理功能。
In order to elucidate the effect of the peroxidase EfeB on the regulation of intracellular oxidative stress in Escherichia coli, Red recombination/disruption and over-expression of the efeB gene were performed. The results showed that after over-expression of efeB, the content of malondialdehyde in the cell was decreased by 85% compared with the parental strain (223.79 nmol/g protein), and the level of intracellular reactive oxygen species was 1.36 folds compared with that of the parental strain. Meanwhile, the expression level of katE was decreased 87.62%, while efeU, efeO, sodA, oxyR and recA up-regulated by 3.88, 1.33, 2.48, 1.95 and 1.49 folds respectively. After exogenous addition of H2O2, katE was further down-regulated by 94.81% while recA was further up-regulated by 8.09 folds in the efeB over-expressing strain Eco/pEE. Conversely, efeB-deficient strain exhibited slower growth rate with higher intracellular malondialdehyde content (1.15-fold of the parental strain). The level of intracellular reactive oxygen species in efeB-deficient strain was 0.5-fold of the parental strain. The results provided new physiological function of EfeB in E.coli under oxidative stress.
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