Department of Biochemical Sciences, CNR Institute of Molecular Biology and Pathology and Istituto Pasteur - Fondazione Cenci Bolognetti, Sapienza University of Rome, I-00185 Rome, Italy.
Biochem Biophys Res Commun. 2010 Sep 3;399(4):654-8. doi: 10.1016/j.bbrc.2010.07.137. Epub 2010 Aug 5.
Flavohemoglobins (flavoHbs), commonly found in bacteria and fungi, afford protection from nitrosative stress by degrading nitric oxide (NO) to nitrate. Giardia intestinalis, a microaerophilic parasite causing one of the most common intestinal human infectious diseases worldwide, is the only pathogenic protozoon as yet identified coding for a flavoHb. By NO amperometry we show that, in the presence of NADH, the recombinant Giardia flavoHb metabolizes NO with high efficacy under aerobic conditions (TN=116+/-10s(-1) at 1microM NO, T=37 degrees C). The activity is [O(2)]-dependent and characterized by an apparent K(M,O2)=22+/-7microM. Immunoblotting analysis shows that the protein is expressed at low levels in the vegetative trophozoites of Giardia; accordingly, these cells aerobically metabolize NO with low efficacy. Interestingly, in response to nitrosative stress (24-h incubation with 5mM nitrite) flavoHb expression is enhanced and the trophozoites thereby become able to metabolize NO efficiently, the activity being sensitive to both cyanide and carbon monoxide. The NO-donors S-nitrosoglutathione (GSNO) and DETA-NONOate mimicked the effect of nitrite on flavoHb expression. We propose that physiologically flavoHb contributes to NO detoxification in G. intestinalis.
Flavohemoglobins (flavoHbs)，常见于细菌和真菌中，通过将一氧化氮 (NO) 降解为硝酸盐来提供抗硝化应激的保护。贾第虫属，一种微需氧寄生虫，是目前唯一被发现编码 flavoHb 的致病性原生动物，可引起全球最常见的肠道人类传染病之一。通过 NO 安培法，我们表明，在 NADH 的存在下，重组的贾第虫属 flavoHb 在有氧条件下以高功效代谢 NO（在 1μM NO 下，T=37°C 时，TN=116+/-10s(-1)）。该活性是[O(2)]依赖性的，并以表观 K(M,O2)=22+/-7μM 为特征。免疫印迹分析表明，该蛋白在贾第虫属的营养体中低水平表达；因此，这些细胞以低功效有氧代谢 NO。有趣的是，在硝化应激（用 5mM 亚硝酸盐孵育 24 小时）下，flavoHb 的表达增强，滋养体因此能够有效地代谢 NO，该活性对氰化物和一氧化碳均敏感。NO 供体 S-亚硝基谷胱甘肽 (GSNO) 和 DETA-NONOate 模拟了亚硝酸盐对 flavoHb 表达的影响。我们提出生理上 flavoHb 有助于 G. intestinalis 中的 NO 解毒。
