Advanced Bio-resource R&D Center, Department of Biology, College of Natural Sciences, Kyungpook National University, #1370 Sankyuk-dong, Buk-gu, Daegu, 702-701, Republic of Korea.
Appl Microbiol Biotechnol. 2013 Oct;97(20):8997-9009. doi: 10.1007/s00253-013-4729-9. Epub 2013 Feb 3.
We investigated Arctic plants to determine if they have a specific mechanism enabling them to adapt to extreme environments because they are subject to such conditions throughout their life cycles. Among the cell defense systems of the Arctic mouse-ear chickweed Cerastium arcticum, we identified a stress-responsive dehydrin gene CaDHN that belongs to the SK5 subclass and contains conserved regions with one S segment at the N-terminus and five K segments from the N-terminus to the C-terminus. To investigate the molecular properties of CaDHN, the yeast Saccharomyces was transformed with CaDHN. CaDHN-expressing transgenic yeast (TG) cells recovered more rapidly from challenge with exogenous stimuli, including oxidants (hydrogen peroxide, menadione, and tert-butyl hydroperoxide), high salinity, freezing and thawing, and metal (Zn(2+)), than wild-type (WT) cells. TG cells were sensitive to copper, cobalt, and sodium dodecyl sulfate. In addition, the cell survival of TG cells was higher than that of WT cells when cells at the mid-log and stationary stages were exposed to increased ethanol concentrations. There was a significant difference in cultures that have an ethanol content >16 %. During glucose-based batch fermentation at generally used (30 °C) and low (18 °C) temperatures, TG cells produced a higher alcohol concentration through improved cell survival. Specifically, the final alcohol concentrations were 13.3 and 13.2 % in TG cells during fermentation at 30 and 18 °C, respectively, whereas they were 10.2 and 9.4 %, respectively, in WT cells under the same fermentation conditions. An in vitro assay revealed that purified CaDHN acted as a reactive oxygen species scavenger by neutralizing H2O2 and a chaperone by preventing high temperature-mediated catalase inactivation. Taken together, our results show that CaDHN expression in transgenic yeast confers tolerance to various abiotic stresses by improving redox homeostasis and enhances fermentation capacity, especially at low temperatures (18 °C).
我们研究了北极植物,以确定它们是否具有特定的机制来适应极端环境,因为它们在整个生命周期中都处于这种条件下。在北极田蓟雀麦 Cerastium arcticum 的细胞防御系统中,我们鉴定出一种应激响应的脱水素基因 CaDHN,它属于 SK5 亚类,包含保守区域,在 N 端有一个 S 片段和从 N 端到 C 端的五个 K 片段。为了研究 CaDHN 的分子特性,我们用 CaDHN 转化酵母 Saccharomyces。与野生型 (WT) 细胞相比,表达 CaDHN 的转基因酵母 (TG) 细胞从包括氧化剂 (过氧化氢、维生素 K3 和叔丁基过氧化氢)、高盐度、冷冻和解冻以及金属 (Zn(2+)) 等外源刺激的挑战中恢复得更快。TG 细胞对铜、钴和十二烷基硫酸钠敏感。此外,当处于对数中期和静止期的细胞暴露于更高的乙醇浓度时,TG 细胞的细胞存活率高于 WT 细胞。在乙醇含量 >16 %的培养物中存在显著差异。在通常使用的 (30°C) 和低温 (18°C) 下进行基于葡萄糖的分批发酵时,TG 细胞通过提高细胞存活率产生了更高的酒精浓度。具体来说,在 30°C 和 18°C 发酵时,TG 细胞的最终酒精浓度分别为 13.3%和 13.2%,而在相同发酵条件下,WT 细胞的最终酒精浓度分别为 10.2%和 9.4%。体外实验表明,纯化的 CaDHN 通过中和 H2O2 作为活性氧物质清除剂,并通过防止高温介导的过氧化氢酶失活作为分子伴侣发挥作用。总之,我们的研究结果表明,在转基因酵母中表达 CaDHN 通过改善氧化还原平衡来赋予对各种非生物胁迫的耐受性,并增强发酵能力,尤其是在低温 (18°C) 下。