Iwadate Yumi, Funabasama Noriyuki, Kato Jun-Ichi
Department of Biological Sciences, Graduate School of Science and Engineering, Tokyo Metropolitan University, 1-1 Minamiohsawa, Hachioji, Tokyo 192-0397, Japan.
FEMS Microbiol Lett. 2017 Nov 1;364(20). doi: 10.1093/femsle/fnx193.
Previously, we constructed a series of reduced-genome strains of Escherichia coli by combining large-scale chromosome deletions and then tested the sensitivity of these strains to the redox-cycling drug menadione. In this study, we analyzed a deletion that increased menadione sensitivity and discovered that loss of selenocysteine synthase genes was responsible for the strain's reduced tolerance to oxidative stress. Mutants of formate dehydrogenases, which are selenocysteine-containing enzymes, were also sensitive to menadione, indicating that these enzymes are involved in oxidative stress during stationary phase, specifically under microaerobic conditions in the presence of glucose. Among three formate dehydrogenases encoded by the E. coli genome, two were responsible for the observed phenotypes: formate dehydrogenase-H and -O. In a mutant of fdhD, which encodes a sulfur transferase that is essential for formate dehydrogenase activity, formate dehydrogenase-O could still contribute to oxidative stress tolerance, revealing a novel role for this protein. Consistent with this, overproduction of the electron transfer subunits of this enzyme, FdoH and FdoI, increased menadione tolerance and supported survival in stationary phase. These results suggested that formate dehydrogenase-O serves as an electron transfer element in glucose metabolism to promote oxidative stress tolerance and survival in stationary phase.
此前，我们通过大规模染色体缺失构建了一系列大肠杆菌的简化基因组菌株，然后测试了这些菌株对氧化还原循环药物甲萘醌的敏感性。在本研究中，我们分析了一个增加甲萘醌敏感性的缺失，发现硒代半胱氨酸合酶基因的缺失是该菌株对氧化应激耐受性降低的原因。甲酸脱氢酶的突变体，即含硒代半胱氨酸的酶，对甲萘醌也敏感，表明这些酶在稳定期参与氧化应激，特别是在存在葡萄糖的微需氧条件下。在大肠杆菌基因组编码的三种甲酸脱氢酶中，有两种导致了观察到的表型：甲酸脱氢酶-H和-O。在fdhD突变体中，fdhD编码对甲酸脱氢酶活性至关重要的硫转移酶，甲酸脱氢酶-O仍可有助于氧化应激耐受性，揭示了该蛋白的新作用。与此一致的是，该酶的电子传递亚基FdoH和FdoI的过量表达增加了甲萘醌耐受性，并支持稳定期的存活。这些结果表明，甲酸脱氢酶-O作为葡萄糖代谢中的电子传递元件，促进氧化应激耐受性和稳定期存活。
