Hu Mimi, Bao Weiwei, Peng Qiqun, Hu Wei, Yang Xinyu, Xiang Yan, Yan Xiongying, Li Mian, Xu Ping, He Qiaoning, Yang Shihui
State Key Laboratory of Biocatalysis and Enzyme Engineering, and School of Life Sciences, Hubei University, Wuhan, China.
Institute of Modern Physics, Chinese Academy of Sciences, Lanzhou, China.
Front Bioeng Biotechnol. 2023 Feb 21;11:1135484. doi: 10.3389/fbioe.2023.1135484. eCollection 2023.
Lactate is the precursor for polylactide. In this study, a lactate producer of was constructed by replacing with gene driven by a strong promoter P, replacing with native gene driven by P and replacing native with another copy of driven by P to divert carbon from ethanol to D-lactate. The resultant strain ZML-pdc-ldh produced 13.8 ± 0.2 g/L lactate and 16.9 ± 0.3 g/L ethanol using 48 g/L glucose. Lactate production of ZML-pdc-ldh was further investigated after fermentation optimization in pH-controlled fermenters. ZML-pdc-ldh produced 24.2 ± 0.6 g/L lactate and 12.9 ± 0.8 g/L ethanol as well as 36.2 ± 1.0 g/L lactate and 40.3 ± 0.3 g/L ethanol, resulting in total carbon conversion rate of 98.3% ± 2.5% and 96.2% ± 0.1% with final product productivity of 1.9 ± 0.0 g/L/h and 2.2 ± 0.0 g/L/h in RMG5 and RMG12, respectively. Moreover, ZML-pdc-ldh produced 32.9 ± 0.1 g/L D-lactate and 27.7 ± 0.2 g/L ethanol as well as 42.8 ± 0.0 g/L D-lactate and 53.1 ± 0.7 g/L ethanol with 97.1% ± 0.0% and 99.1% ± 0.8% carbon conversion rate using 20% molasses or corncob residue hydrolysate, respectively. Our study thus demonstrated that it is effective for lactate production by fermentation condition optimization and metabolic engineering to strengthen heterologous expression while reducing the native ethanol production pathway. The capability of recombinant lactate-producer of for efficient waste feedstock conversion makes it a promising biorefinery platform for carbon-neutral biochemical production.
乳酸是聚乳酸的前体。在本研究中，通过用强启动子P驱动的基因替换基因、用P驱动的天然基因替换基因以及用P驱动的另一个拷贝替换天然基因，将碳从乙醇转移到D - 乳酸，构建了一株乳酸生产菌。所得菌株ZML - pdc - ldh以48 g/L葡萄糖为原料，产生13.8±0.2 g/L乳酸和16.9±0.3 g/L乙醇。在pH控制的发酵罐中进行发酵优化后，进一步研究了ZML - pdc - ldh的乳酸生产情况。ZML - pdc - ldh分别在RMG5和RMG12中产生24.2±0.6 g/L乳酸和12.9±0.8 g/L乙醇以及36.2±1.0 g/L乳酸和40.3±0.3 g/L乙醇，总碳转化率分别为98.3%±2.5%和96.2%±0.1%，最终产物生产率分别为1.9±0.0 g/(L·h)和2.2±0.0 g/(L·h)。此外，ZML - pdc - ldh分别使用20%糖蜜或玉米芯残渣水解液，产生32.9±0.1 g/L D - 乳酸和27.7±之.2 g/L乙醇以及42.8±0.0 g/L D - 乳酸和53.1±0.7 g/L乙醇，碳转化率分别为97.1%±0.0%和99.1%±0.8%。因此，我们的研究表明，通过优化发酵条件和代谢工程，在减少天然乙醇生产途径的同时加强异源表达，对于乳酸生产是有效的。重组乳酸生产菌高效转化废弃原料的能力使其成为一个有前途的用于碳中和生物化学产品生产的生物精炼平台。
