Korea Institute of Ocean Science and Technology, Ansan, 15627, Republic of Korea.
Biomass and Waste Energy Laboratory, Korea Institute of Energy Research, Daejeon, 34129, Republic of Korea.
Appl Microbiol Biotechnol. 2017 Jun;101(12):5081-5088. doi: 10.1007/s00253-017-8234-4. Epub 2017 Mar 24.
The F-reducing hydrogenase of methanogens functions in methanogenesis by providing reduced coenzyme F (FH) as an electron donor. In non-methanogens, however, their physiological function has not been identified yet. In this study, we constructed an ΔfrhA mutant, whose frhA gene encoding the hydrogenase α subunit was deleted, in the non-methanogenic Thermococcus onnurineus NA1 as a model organism. There was no significant difference in the formate-dependent growth between the mutant and the wild-type strains. Interestingly, the mutation in the frhA gene affected the expression of genes involved in various cellular functions such as H oxidation, chemotactic signal transduction, and carbon monoxide (CO) metabolism. Among these genes, the CO oxidation gene cluster, enabling CO-dependent growth and H production, showed a 2.8- to 7.0-fold upregulation by microarray-based whole transcriptome expression profiling. The levels of proteins produced by this gene cluster were also significantly increased not only under the formate condition but also under the CO condition. In a controlled bioreactor, where 100% CO was continuously fed, the ΔfrhA mutant exhibited significant increases in cell growth (2.8-fold) and H production (3.4-fold). These findings strongly imply that this hydrogenase is functional in non-methanogens and is related to various cellular metabolic processes through an unidentified mechanism. An understanding of the mechanism by which the frhA gene deletion affected the expression of other genes will provide insights that can be applied to the development of strategies for the enhancement of H production using CO as a substrate.
产甲烷菌的 F 型氢化酶通过提供还原型辅酶 F（FH）作为电子供体在产甲烷作用中发挥作用。然而，在非产甲烷菌中，其生理功能尚未确定。在这项研究中，我们构建了一个ΔfrhA 突变体，其 frhA 基因编码氢化酶α亚基缺失，作为模型生物的非产甲烷古菌 Thermococcus onnurineus NA1。突变体和野生型菌株在依赖甲酸盐的生长方面没有显著差异。有趣的是，frhA 基因的突变影响了涉及各种细胞功能的基因的表达，如 H 氧化、趋化信号转导和一氧化碳（CO）代谢。在这些基因中，CO 氧化基因簇能够进行 CO 依赖性生长和 H 生产，通过基于微阵列的全转录组表达谱分析显示其表达上调了 2.8 到 7.0 倍。该基因簇产生的蛋白质水平不仅在甲酸盐条件下，而且在 CO 条件下也显著增加。在一个连续进料 100%CO 的控制生物反应器中，ΔfrhA 突变体的细胞生长（2.8 倍）和 H 生产（3.4 倍）显著增加。这些发现强烈表明，这种氢化酶在非产甲烷菌中是有功能的，并且通过一种未知的机制与各种细胞代谢过程有关。了解 frhA 基因缺失如何影响其他基因的表达的机制将为利用 CO 作为底物增强 H 生产的策略的开发提供有价值的见解。
