Isolation of cobalt hyper-resistant mutants of Saccharomyces cerevisiae by in vivo evolutionary engineering approach
Isolation of cobalt hyper-resistant mutants of Saccharomyces cerevisiae by in vivo evolutionary engineering approach
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DOI：
10.1016/j.jbiotec.2009.06.024
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发表时间：
2009-08-20
影响因子：
4.1
通讯作者：
Francois, Jean M.
中科院分区：
文献类型：
--
作者：
Cakar, Z. Petek;Alkim, Ceren;Francois, Jean M.
钴是一种具有磁性的重要元素，用于各种工业应用。但也是生物活性所必需的。关于生命系统对钴应激的细胞反应知之甚少。为了研究这一机制，我们分离了对高达 8 mmol l(-1) 的高浓度钴具有抗性的酿酒酵母细胞。通过采用四种不同的“体内”进化工程策略：在恒定或逐渐增加的压力水平下进行选择，以及在连续或脉冲暴露于钴压力下进行选择。在持续暴露于逐渐增加的钴胁迫水平下的选择产生了对钴最具抵抗力的细胞群。然而，在突变群体中，耐药性是高度异质性的，在最耐药群体中，孤立个体对8 mmol l(-1) CoCl(2) 的存活率是3 至3700 倍。此外，与野生型相比，抗钴单个菌落的细胞内钴含量低2-4倍，并且对镍、锌、锰等金属具有交叉抗性，但对铜和铬离子没有交叉抗性。与连续暴露于钴下进化的突变体相反，通过脉冲暴露策略分离的突变体也表现出对热休克和过氧化氢应激的抵抗力。总而言之，这项研究强化了这样一个事实，即进化工程可用于选择具有非常特定表型的菌株，并进一步说明了选择分离最佳进化菌株的策略的重要性。 (C) 2009 Elsevier B.V. 保留所有权利。
Cobalt is an important element with magnetic properties used in various industrial applications. but is also needed for biological activity. Very little is known about the cellular response of living systems to cobalt stress. Towards investigating this mechanism, we isolated individual Saccharomyces cerevisiae cells resistant to high cobalt concentrations up to 8 mmol l(-1). by employing four different 'in vivo' evolutionary engineering strategies: selection under constant or gradually increasing stress levels, and selection under continuous or pulse exposure to cobalt stress. Selection under continuous exposure to gradually increasing cobalt stress levels yielded the most resistant cell population to cobalt. However, the resistance was highly heterogeneous within the mutant populations ranging from 3- to 3700-fold survival rate of isolated individuals to 8 mmol l(-1) CoCl(2) in the most resistant population. Moreover, cobalt-resistant individual colonies were associated with 2-4-times lower intracellular cobalt contents as compared to wild-type, and with cross-resistance to metals such as nickel, zinc, manganese, but not to copper and chromium ions. Contrary to mutants evolved under continuous exposure to cobalt, those isolated by pulse exposure strategy also exhibited resistance to heat shock and hydrogen peroxide stress. Taken together, this study reinforced the fact that evolutionary engineering is useful in selecting strains with very specific phenotypes, and further illustrated the importance of the strategy chosen to isolate the best evolved strain. (C) 2009 Elsevier B.V. All rights reserved.