Institute of Microbial Technology, Sector 39 A, Chandigarh 160036, India.
Biochem J. 2010 Feb 24;426(3):271-80. doi: 10.1042/BJ20091417.
The bacterial haemoglobin from Vitreoscilla, VHb, displays several unusual properties that are unique among the globin family. When the gene encoding VHb, vgb, is expressed from its natural promoter in either Vitreoscilla or Escherichia coli, the level of VHb increases more than 50-fold under hypoxic conditions and decreases significantly during oxidative stress, suggesting similar functioning of the vgb promoter in both organisms. In the present study we show that expression of VHb in E. coli induced the antioxidant genes katG (catalase-peroxidase G) and sodA (superoxide dismutase A) and conferred significant protection from oxidative stress. In contrast, when vgb was expressed in an oxyR mutant of E. coli, VHb levels increased and the strain showed high sensitivity to oxidative stress without induction of antioxidant genes; this indicates the involvement of the oxidative stress regulator OxyR in mediating the protective effect of VHb under oxidative stress. A putative OxyR-binding site was identified within the vgb promoter and a gel-shift assay confirmed its interaction with oxidized OxyR, an interaction which was disrupted by the reduced form of the transcriptional activator Fnr (fumurate and nitrate reductase). This suggested that the redox state of OxyR and Fnr modulates their interaction with the vgb promoter. VHb associated with reduced OxyR in two-hybrid screen experiments and in vitro, converting it into an oxidized state in the presence of NADH, a condition where VHb is known to generate H2O2. These observations unveil a novel mechanism by which VHb may transmit signals to OxyR to autoregulate its own biosynthesis, simultaneously activating oxidative stress functions. The activation of OxyR via VHb, reported in the present paper for the first time, suggests the involvement of VHb in transcriptional control of many other genes as well.
来自威氏红球菌的细菌血红蛋白 VHb 具有几种在球蛋白家族中独一无二的不寻常特性。当编码 VHb 的基因 vgb 从其天然启动子在威氏红球菌或大肠杆菌中表达时，在低氧条件下 VHb 的水平会增加 50 多倍，而在氧化应激期间则会显著下降，这表明 vgb 启动子在这两种生物体中的功能相似。在本研究中，我们表明在大肠杆菌中表达 VHb 会诱导抗氧化基因 katG（过氧化氢酶过氧化物酶 G）和 sodA（超氧化物歧化酶 A）的表达，并赋予其对氧化应激的显著保护作用。相比之下，当 vgb 在大肠杆菌的 oxyR 突变体中表达时，VHb 水平增加，且该菌株在没有诱导抗氧化基因的情况下对氧化应激高度敏感；这表明氧化应激调节剂 OxyR 参与介导 VHb 在氧化应激下的保护作用。在 vgb 启动子内鉴定出一个推定的 OxyR 结合位点，凝胶迁移实验证实了其与氧化 OxyR 的相互作用，而该相互作用被转录激活剂 Fnr（延胡索酸和硝酸盐还原酶）的还原形式破坏。这表明 OxyR 和 Fnr 的氧化还原状态调节它们与 vgb 启动子的相互作用。在双杂交筛选实验中和体外，VHb 与还原型 OxyR 相关联，在存在 NADH 的情况下将其转化为氧化状态，在这种情况下，已知 VHb 会产生 H2O2。这些观察结果揭示了一种新的机制，通过该机制，VHb 可以将信号传递给 OxyR 来自我调节其自身的生物合成，同时激活氧化应激功能。本文首次报道了通过 VHb 激活 OxyR，这表明 VHb 可能参与了许多其他基因的转录控制。
