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用于检测耐辐射球菌中活性氧清除活性及对DNA氧化损伤抑制作用的化学发光测定法。

Chemiluminescence assay for reactive oxygen species scavenging activities and inhibition on oxidative damage of DNA in Deinococcus radiodurans.

作者信息

Tian Bing, Wu Yuanyuan, Sheng Duohong, Zheng Zhiguo, Gao Guanjun, Hua Yuejin

机构信息

Institute of Nuclear-Agricultural Sciences, Zhejiang University, 310029 Hangzhou, China.

出版信息

Luminescence. 2004 Mar-Apr;19(2):78-84. doi: 10.1002/bio.761.

Abstract

Free radical scavenging effects of the cellular protein extracts from two strains of Deinococcus radiodurans and Escherichia coli against O2-, H2O2 and *OH were investigated by chemiluminescence (CL) methods. The cellular protein extracts of D. radiodurans R1 and KD8301 showed higher scavenging effects on O2- than that of E. coli. D. radiodurans R1 and KD8301 also strongly scavenged H2O2 with an EC50 (50% effective concentration) of 0.12 and 0.2 mg/mL, respectively, compared to that of E. coli (EC50 = 3.56 mg/mL). The two strains of D. radiodurans were effective in scavenging *OH generated by the Fenton reaction, with EC50 of 0.059 and 0.1 mg/mL, respectively, compared to that of E. coli (EC50 > 1 mg/mL). Results from the chemiluminescence assay of *OH-induced DNA damage and the plasmid pUC18 DNA double-strand break (DSB) model in vitro showed that D. radiodurans had remarkably inhibitory effect on the *OH-induced oxidative damage of DNA. The scavenging effects of D. radiodurans on reactive oxygen species (ROS) played an important role in the response to oxidation stress and preventing against DNA oxidative damage, and may be attributed to intracellular scavenging proteins, including superoxide dismutase (SOD) and catalase.

摘要

采用化学发光(CL)方法研究了两株耐辐射球菌(Deinococcus radiodurans)和大肠杆菌(Escherichia coli)细胞蛋白提取物对超氧阴离子(O2-)、过氧化氢(H2O2)和羟基自由基(·OH)的自由基清除作用。耐辐射球菌R1和KD8301的细胞蛋白提取物对O2-的清除作用高于大肠杆菌。与大肠杆菌(EC50 = 3.56 mg/mL)相比,耐辐射球菌R1和KD8301对H2O2也有很强的清除作用(EC50分别为0.12和0.2 mg/mL)。两株耐辐射球菌对芬顿反应产生的·OH有清除作用,EC50分别为0.059和0.1 mg/mL,而大肠杆菌的EC50>1 mg/mL。体外·OH诱导的DNA损伤化学发光分析和质粒pUC18 DNA双链断裂(DSB)模型结果表明,耐辐射球菌对·OH诱导的DNA氧化损伤有显著抑制作用。耐辐射球菌对活性氧(ROS)的清除作用在其对氧化应激的反应和防止DNA氧化损伤中起重要作用,这可能归因于细胞内的清除蛋白,包括超氧化物歧化酶(SOD)和过氧化氢酶。

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