Wildschut Janine D, Lang R Michael, Voordouw Johanna K, Voordouw Gerrit
Department of Biological Sciences, University of Calgary, 2500 University Drive NW, Calgary, Alberta T2N 1N4, Canada.
J Bacteriol. 2006 Sep;188(17):6253-60. doi: 10.1128/JB.00425-06.
Genes for superoxide reductase (Sor), rubredoxin (Rub), and rubredoxin:oxygen oxidoreductase (Roo) are located in close proximity in the chromosome of Desulfovibrio vulgaris Hildenborough. Protein blots confirmed the absence of Roo from roo mutant and sor-rub-roo (srr) mutant cells and its presence in sor mutant and wild-type cells grown under anaerobic conditions. Oxygen reduction rates of the roo and srr mutants were 20 to 40% lower than those of the wild type and the sor mutant, indicating that Roo functions as an O2 reductase in vivo. Survival of single cells incubated for 5 days on agar plates under microaerophilic conditions (1% air) was 85% for the sor, 4% for the roo, and 0.7% for the srr mutant relative to that of the wild type (100%). The similar survival rates of sor mutant and wild-type cells suggest that O2 reduction by Roo prevents the formation of reactive oxygen species (ROS) under these conditions; i.e., the ROS-reducing enzyme Sor is only needed for survival when Roo is missing. In contrast, the sor mutant was inactivated much more rapidly than the roo mutant when liquid cultures were incubated in 100% air, indicating that O2 reduction by Roo and other terminal oxidases did not prevent ROS formation under these conditions. Competition of Sor and Roo for limited reduced Rub was suggested by the observation that the roo mutant survived better than the wild type under fully aerobic conditions. The roo mutant was more strongly inhibited than the wild type by the nitric oxide (NO)-generating compound S-nitrosoglutathione, indicating that Roo may also serve as an NO reductase in vivo.
超氧化物还原酶（Sor）、红素氧还蛋白（Rub）和红素氧还蛋白：氧氧化还原酶（Roo）的基因在嗜热栖热脱硫弧菌（Desulfovibrio vulgaris Hildenborough）的染色体中紧密相邻。蛋白质印迹证实，在roo突变体和sor-rub-roo（srr）突变体细胞中不存在Roo，而在厌氧条件下生长的sor突变体和野生型细胞中存在Roo。roo和srr突变体的氧还原率比野生型和sor突变体低20%至40%，表明Roo在体内作为一种O2还原酶发挥作用。在微需氧条件（1%空气）下，在琼脂平板上培养5天的单细胞存活率相对于野生型（100%）而言，sor突变体为85%，roo突变体为4%，srr突变体为0.7%。sor突变体和野生型细胞相似的存活率表明，在这些条件下，Roo对O2的还原可防止活性氧（ROS）的形成；即，只有当Roo缺失时，ROS还原酶Sor才是生存所必需的。相比之下，当液体培养物在100%空气中孵育时，sor突变体比roo突变体更快失活，这表明在这些条件下，Roo和其他末端氧化酶对O2的还原并不能防止ROS的形成。在完全好氧条件下，roo突变体比野生型存活得更好，这一观察结果表明Sor和Roo对有限的还原型Rub存在竞争。一氧化氮（NO）生成化合物S-亚硝基谷胱甘肽对roo突变体的抑制作用比野生型更强，这表明Roo在体内也可能作为一种NO还原酶发挥作用。
