Bryant C, DeLuca M
Department of Chemistry, University of California, San Diego, La Jolla 92093.
J Biol Chem. 1991 Mar 5;266(7):4119-25.
The reductive products of several nitroaromatic compounds have been found to be toxic, mutagenic, and carcinogenic. The nitroreductases present in intestinal microflora have been implicated in the biotransformation of these compounds to their deleterious metabolites. A "classical" nitroreductase has been purified from Enterobacter cloacae 587-fold using a protocol which yields approximately 1 mg of purified nitroreductase from 10 liters of cell culture. An analysis of the physical properties of the nitroreductase indicates that the enzyme is active as a monomer with a calculated molecular mass of 27 kDa. FMN has been identified as a required flavin cofactor and is present at a stoichiometry of 0.88 mol of FMN bound/mol of active enzyme. The enzyme was found capable of reducing nitrofurazone under aerobic conditions indicating that the mechanism involves an obligatory two-electron transfer. Thus, this enzyme can be classified as an oxygen-insensitive nitroreductase. The purified nitroreductase can utilize either NADH or NADPH as a source of reducing equivalents and can reduce a variety of nitroaromatic compounds including nitrofurans and nitrobenzenes as well as quinones. Studies in which the rates of nitroreduction for a series of para substituted nitrobenzene derivatives were determined suggest that a linear free energy relationship exists between the rate and the redox midpoint potential of the substrate.
已发现几种硝基芳香化合物的还原产物具有毒性、致突变性和致癌性。肠道微生物群中存在的硝基还原酶与这些化合物生物转化为其有害代谢物有关。一种“经典”硝基还原酶已从阴沟肠杆菌中纯化出来，采用的方案可从10升细胞培养物中获得约1毫克纯化的硝基还原酶，纯化倍数为587倍。对硝基还原酶物理性质的分析表明，该酶以单体形式具有活性，计算分子量为27 kDa。已确定FMN是必需的黄素辅因子，其化学计量比为每摩尔活性酶结合0.88摩尔FMN。发现该酶能够在有氧条件下还原呋喃西林，这表明其机制涉及强制性的双电子转移。因此，这种酶可归类为对氧不敏感的硝基还原酶。纯化的硝基还原酶可以利用NADH或NADPH作为还原当量的来源，并且可以还原多种硝基芳香化合物，包括硝基呋喃和硝基苯以及醌。对一系列对位取代硝基苯衍生物的硝基还原速率进行测定的研究表明，底物的还原速率与氧化还原中点电位之间存在线性自由能关系。
