ISOLATION OF SUPEROXIDE-DISMUTASE MUTANTS IN ESCHERICHIA-COLI - IS SUPEROXIDE-DISMUTASE NECESSARY FOR AEROBIC LIFE
ISOLATION OF SUPEROXIDE-DISMUTASE MUTANTS IN ESCHERICHIA-COLI - IS SUPEROXIDE-DISMUTASE NECESSARY FOR AEROBIC LIFE
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DOI：
10.1002/j.1460-2075.1986.tb04256.x
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发表时间：
1986-03-01
期刊：
影响因子：
11.4
通讯作者：
TOUATI, D
中科院分区：
文献类型：
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作者：
CARLIOZ, A;TOUATI, D
携带氯霉素抗性标记的Mu转座子已被插入克隆的大肠杆菌基因sodA和sodB中，分别编码锰超氧化物歧化酶（MnSOD）和铁超氧化物歧化酶（FeSOD），从而产生突变和基因融合。通过等位基因交换将突变的 sodA 或 sodB 基因引入细菌染色体中。通过活性测量和免疫印迹分析，所得突变体缺乏相应的 SOD。在需氧条件下，在丰富的培养基中，缺乏 FeSOD 或 MnSOD 对生长或对超氧化物生成剂百草枯的敏感性没有重大影响。在基本培养基中，需氧生长未受影响，但对百草枯的敏感性增加，尤其是在 sodA 突变体中。还获得了完全不含SOD的sodA sodB双突变体。它能够在丰富的培养基中有氧生长，其过氧化氢酶水平不受影响，并且对百草枯和过氧化氢高度敏感；双突变体无法在基本葡萄糖培养基上有氧生长。可以通过去除氧气、提供 SOD 过量产生的质粒或通过向培养基补充 20 种氨基酸来恢复生长。结论是，大肠杆菌中完全缺乏 SOD 会产生对氧的条件敏感性。
Mu transposons carrying the chloramphenicol resistance marker have been inserted into the cloned Escherichia coli genes sodA and sodB coding for manganese superoxide dismutase (MnSOD) and iron superoxide dismutase (FeSOD) respectively, creating mutations and gene fusions. The mutated sodA or sodB genes were introduced into the bacterial chromosome by allelic exchange. The resulting mutants were shown to lack the corresponding SOD by activity measurement and immunoblot analysis. Aerobically, in rich medium, the absence of FeSOD or MnSOD had no major effect on growth or sensitivity to the superoxide generator, paraquat. In minimal medium aerobic growth was not affected, but the sensitivity to paraquat was increased, especially in the sodA mutant. A sodA sodB double mutant completely devoid of SOD was also obtained. It was able to grow aerobically in rich medium, its catalase level was unaffected and it was highly sensitive to paraquat and hydrogen peroxide; the double mutant was unable to grow aerobically on minimal glucose medium. Growth could be restored by removing oxygen, by providing an SOD-overproducing plasmid or by supplementing the medium with the 20 amino acids. It is concluded that the total absence of SOD in E. coli creates a conditional sensitivity to oxygen.