Jacobson F S, Morgan R W, Christman M F, Ames B N
Department of Biochemistry, University of California, Berkeley 94720.
J Biol Chem. 1989 Jan 25;264(3):1488-96.
A peroxide reductase (peroxidase) which converts lipid hydroperoxides and other alkyl hydroperoxides to the corresponding alcohols, using either NADH or NADPH as the reducing agent, has been identified in both Salmonella typhimurium and Escherichia coli. This enzyme is shown to play a role in protecting against alkyl hydroperoxide mutagenesis. To our knowledge this work represents the first description of an NAD(P)H peroxidase in enteric bacteria and the first reported bacterial peroxidase to exhibit high activity toward alkyl hydroperoxides. A high performance liquid chromatography-based assay for the alkyl hydroperoxide reductase has been developed by monitoring the reduction of cumene hydroperoxide, a model alkyl hydroperoxide. By using this assay, the enzyme has been purified from a S. typhimurium regulatory mutant, oxyR1, which overexpresses a number of proteins involved in defenses against oxidative damage, and which contains 20-fold more of the alkyl hydroperoxide reductase than the wild-type strain. The purified activity requires the presence of two separable components having subunit molecular weights of 22,000 and 57,000. The 57-kDa protein contains a bound FAD cofactor and can use either NADH or NADPH as an electron donor for the direct reduction of redox dyes, or of alkyl hydroperoxides when combined with the 22-kDa protein. This enzyme may thus serve as a prokaryotic equivalent to the glutathione reductase/glutathione peroxidase system in eukaryotes.
在鼠伤寒沙门氏菌和大肠杆菌中均已鉴定出一种过氧化物还原酶（过氧化物酶），该酶可利用NADH或NADPH作为还原剂，将脂质氢过氧化物和其他烷基氢过氧化物转化为相应的醇。已证明这种酶在防止烷基氢过氧化物诱变中发挥作用。据我们所知，这项工作首次描述了肠道细菌中的NAD(P)H过氧化物酶，也是首次报道的对烷基氢过氧化物具有高活性的细菌过氧化物酶。通过监测异丙苯过氧化氢（一种典型的烷基氢过氧化物）的还原反应，开发了一种基于高效液相色谱的烷基氢过氧化物还原酶检测方法。利用该检测方法，已从鼠伤寒沙门氏菌调节突变体oxyR1中纯化出该酶，该突变体过表达多种参与抗氧化损伤防御的蛋白质，其烷基氢过氧化物还原酶的含量比野生型菌株高20倍。纯化后的活性需要两种可分离的组分，其亚基分子量分别为22,000和57,000。57 kDa的蛋白质含有结合的FAD辅因子，可使用NADH或NADPH作为电子供体，直接还原氧化还原染料，或与22 kDa的蛋白质结合时还原烷基氢过氧化物。因此，这种酶可能相当于真核生物中的谷胱甘肽还原酶/谷胱甘肽过氧化物酶系统。
