Improving phloroglucinol tolerance and production in Escherichia coli by GroESL overexpression.
Improving phloroglucinol tolerance and production in Escherichia coli by GroESL overexpression.
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通过 GroESL 过表达提高大肠杆菌的间苯三酚耐受性和产量
DOI：
10.1186/s12934-017-0839-x
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发表时间：
2017-12-19
影响因子：
6.4
通讯作者：
Liu H
中科院分区：
文献类型：
--
作者：
Zhang R;Cao Y;Liu W;Xian M;Liu H
间苯三酚是利用工程大肠杆菌成功生产的一种重要化学物质。然而，间苯三酚的毒性会极大地抑制大肠杆菌细胞的生长和活力，而且其生产率仍然太低，不适合工业应用。因此，菌株对有毒代谢物的耐受性仍然是使用生物过程生产化学品的关键问题。在本研究中，我们研究了不同过表达水平的天然GroESL伴侣系统对大肠杆菌间苯三酚耐受性和产量的影响。将groESL基因克隆到表达载体中，其表达水平受三种不同启动子（natural、tac和T7启动子）调控。采用活细胞计数和间苯三酚产量评估菌株耐受性。与对照菌株相比，所有过表达菌株在细胞活力和间苯三酚合成方面均表现出良好的特征。结果表明，在tac启动子下过表达GroESL的菌株在所有试验中表现出最好的耐受性，在0.7 g/L间苯三酚的条件下，其活菌数比琼脂平板培养的对照菌株增加了3.19倍，分批补料发酵下的间苯三酚产量增加了39.5%。该工程菌株在诱导后10 h的补料分批培养中积累间苯三酚达5.3 g/L，产量为0.53 g/L/h。到目前为止，与以前的报道相比，本研究取得了最高的间苯三酚产量，这有望使生物工艺从经济角度上可行。结果表明，适宜的GroESL表达水平在提高大肠杆菌间苯三酚耐受性和产量中起着关键作用，可能通过上调GroESL参与调控胁迫反应系统的某些方面。因此，GroESL过表达是提高大肠杆菌耐受性的一种可行而有效的方法。本文的在线版本（10.1186/s12934-017-0839-x）包含补充材料，授权用户可以使用。
Phloroglucinol is an important chemical which has been successfully produced by engineered Escherichia coli. However, the toxicity of phloroglucinol can enormously inhibit E. coli cell growth and viability, and the productivity is still too low and not economically feasible for industrial applications. Therefore, strain tolerance to toxic metabolites remains a key issue during the production of chemicals using biological processes. In the present work, we examined the impact of the native GroESL chaperone system with different overexpression levels on phloroglucinol tolerance and production in E. coli. The groESL gene was cloned into an expression vector, of which expression level was regulated by three different promoters (natural, tac and T7 promoter). Strain tolerance was evaluated employing viable cell counts and phloroglucinol production. In comparison with the control strain, all GroESL overexpressing strains showed good characteristics in cell viability and phloroglucinol synthesis. Strain which overexpressed GroESL under tac promoter was found to show the best tolerance in all of those tested, resulting in a 3.19-fold increase in viable cell numbers compared with control strain of agar-plate culture under the condition of 0.7 g/L phloroglucinol, and a 39.5% increase in phloroglucinol production under fed-batch fermentation. This engineered strain finally accumulated phloroglucinol up to 5.3 g/L in the fed-batch cultivation 10 h after induction, and the productivity was 0.53 g/L/h. To date, the highest phloroglucinol production was achieved in this work compared with the previous reports, which is promising to make the bioprocess feasible from the economical point. The data show that appropriate expression level of GroESL plays a critical role in improving phloroglucinol tolerance and production in E. coli, and maybe involve in controlling some aspects of the stress response system through upregulation of GroESL. GroESL overexpression is therefore a feasible and efficient approach for improvement of E. coli tolerance. The online version of this article (10.1186/s12934-017-0839-x) contains supplementary material, which is available to authorized users.
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影响因子：
14.8
作者：
de Marco, Ario
通讯作者：
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影响因子：
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作者：
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影响因子：
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影响因子：
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