Lab of Biorefinery, Shanghai Advanced Research Institute, Chinese Academy of Sciences, No. 99 Haike Road, Pudong, Shanghai 201210, PR China; University of Chinese Academy of Sciences, Beijing 100049, PR China.
Josip Juraj Strossmayer University of Osijek, Faculty of Food Technology Osijek, Franje Kuhača 18, Osijek HR-31000, Croatia.
Enzyme Microb Technol. 2024 Jun;177:110438. doi: 10.1016/j.enzmictec.2024.110438. Epub 2024 Mar 19.
Klebsiella pneumoniae can use glucose or glycerol as carbon sources to produce 1,3-propanediol or 2,3-butanediol, respectively. In the metabolism of Klebsiella pneumoniae, hydrogenase-3 is responsible for H production from formic acid, but it is not directly related to the synthesis pathways for 1,3-propanediol and 2,3-butanediol. In the first part of this research, hycEFG, which encodes subunits of the enzyme hydrogenase-3, was knocked out, so K. pneumoniae ΔhycEFG lost the ability to produce H during cultivation using glycerol as a carbon source. As a consequence, the concentration of 1,3-propanediol increased and the substrate (glycerol) conversion ratio reached 0.587 mol/mol. Then, K. pneumoniae ΔldhAΔhycEFG was constructed to erase lactic acid synthesis which led to the further increase of 1,3-propanediol concentration. A substrate (glycerol) conversion ratio of 0.628 mol/mol in batch conditions was achieved, which was higher compared to the wild type strain (0.545 mol/mol). Furthermore, since adhE encodes an alcohol dehydrogenase that catalyzes ethanol production from acetaldehyde, K. pneumoniae ΔldhAΔadhEΔhycEFG was constructed to prevent ethanol production. Contrary to expectations, this did not lead to a further increase, but to a decrease in 1,3-propanediol production. In the second part of this research, glucose was used as the carbon source to produce 2,3-butanediol. Knocking out hycEFG had distinct positive effect on 2,3-butanediol production. Especially in K. pneumoniae ΔldhAΔadhEΔhycEFG, a substrate (glucose) conversion ratio of 0.730 mol/mol was reached, which is higher compared to wild type strain (0.504 mol/mol). This work suggests that the inactivation of hydrogenase-3 may have a global effect on the metabolic regulation of K. pneumoniae, leading to the improvement of the production of two industrially important bulk chemicals, 1,3-propanediol and 2,3-butanediol.
肺炎克雷伯氏菌可以使用葡萄糖或甘油作为碳源，分别生产 1,3-丙二醇或 2,3-丁二醇。在肺炎克雷伯氏菌的代谢过程中，氢化酶-3负责从甲酸中产生 H，但它与 1,3-丙二醇和 2,3-丁二醇的合成途径没有直接关系。在本研究的第一部分中，敲除了编码氢化酶-3亚基的 hycEFG，因此 K. pneumoniae ΔhycEFG 在以甘油为碳源培养时丧失了产生 H 的能力。结果，1,3-丙二醇的浓度增加，底物（甘油）转化率达到 0.587 mol/mol。然后，构建了 K. pneumoniae ΔldhAΔhycEFG 以消除乳酸合成，从而进一步提高 1,3-丙二醇的浓度。在批处理条件下实现了 0.628 mol/mol 的底物（甘油）转化率，与野生型菌株（0.545 mol/mol）相比有所提高。此外，由于 adhE 编码一种醇脱氢酶，可催化乙醛转化为乙醇，因此构建了 K. pneumoniae ΔldhAΔadhEΔhycEFG 以防止乙醇生成。出乎意料的是，这并没有导致 1,3-丙二醇产量的进一步提高，反而导致其产量下降。在本研究的第二部分中，使用葡萄糖作为碳源生产 2,3-丁二醇。敲除 hycEFG 对 2,3-丁二醇的生产有明显的积极影响。特别是在 K. pneumoniae ΔldhAΔadhEΔhycEFG 中，达到了 0.730 mol/mol 的底物（葡萄糖）转化率，与野生型菌株（0.504 mol/mol）相比有所提高。这项工作表明，氢化酶-3 的失活可能对肺炎克雷伯氏菌的代谢调控产生全局影响，从而提高两种重要的工业大宗化学品，1,3-丙二醇和 2,3-丁二醇的产量。
