Hon Shuen, Holwerda Evert K, Worthen Robert S, Maloney Marybeth I, Tian Liang, Cui Jingxuan, Lin Paul P, Lynd Lee R, Olson Daniel G
1Thayer School of Engineering, Dartmouth College, 14 Engineering Drive, Hanover, NH 03755 USA.
Bioenergy Science Center, Oak Ridge National Laboratories, Oak Ridge, TN 37830 USA.
Biotechnol Biofuels. 2018 Sep 6;11:242. doi: 10.1186/s13068-018-1245-2. eCollection 2018.
has been the subject of multiple metabolic engineering strategies to improve its ability to ferment cellulose to ethanol, with varying degrees of success. For ethanol production in , the conversion of pyruvate to acetyl-CoA is catalyzed primarily by the pyruvate ferredoxin oxidoreductase (PFOR) pathway. , which was previously engineered to produce ethanol of high yield (> 80%) and titer (70 g/L), also uses a pyruvate ferredoxin oxidoreductase, , for ethanol production.
Here, we introduced the and ferredoxin into . The introduction of resulted in significant improvements to ethanol yield and titer in grown on 50 g/L of cellobiose, but only when four other genes (, , , and ) were also present. ferredoxin did not have any observable impact on ethanol production. The improvement to ethanol production was sustained even when all annotated native genes were deleted. On high cellulose concentrations, the maximum ethanol titer achieved by this engineered strain from 100 g/L Avicel was 25 g/L, compared to 22 g/L for the reference strain, LL1319 (()-()- ()) under similar conditions. In addition, we also observed that deletion of the results in a significant decrease in isobutanol production.
Here, we demonstrate that the gene can improve ethanol production in as part of the pyruvate-to-ethanol pathway. In our previous strain, high-yield (~ 75% of theoretical) ethanol production could be achieved with at most 20 g/L substrate. In this strain, high-yield ethanol production can be achieved up to 50 g/L substrate. Furthermore, the introduction of increased the maximum titer by 14%.
为提高其将纤维素发酵为乙醇的能力，已开展了多种代谢工程策略研究，取得了不同程度的成功。在[具体生物名称]中生产乙醇时，丙酮酸向乙酰辅酶A的转化主要由丙酮酸铁氧化还原酶（PFOR）途径催化。[具体生物名称]先前经工程改造可实现高产率（>80%）和高滴度（70 g/L）乙醇生产，其乙醇生产也使用丙酮酸铁氧化还原酶[具体酶名称]。
在此，我们将[具体基因名称]和铁氧化还原蛋白引入[具体生物名称]。引入[具体基因名称]后，在以50 g/L纤维二糖为生长底物的[具体生物名称]中，乙醇产率和滴度显著提高，但前提是同时存在其他四个[具体生物名称]基因（[基因名称1]、[基因名称2]、[基因名称3]和[基因名称4]）。铁氧化还原蛋白对乙醇生产没有任何可观察到的影响。即使删除了所有注释的天然[具体生物名称]基因，乙醇生产的改善仍得以维持。在高纤维素浓度下，该工程改造的[具体生物名称]菌株从100 g/L微晶纤维素获得的最大乙醇滴度为25 g/L，而在类似条件下，参考菌株LL1319（[菌株相关描述]）为22 g/L。此外，我们还观察到删除[具体基因名称]会导致异丁醇产量显著下降。
在此，我们证明[具体基因名称]作为[具体生物名称]丙酮酸到乙醇途径的一部分，可提高[具体生物名称]中的乙醇产量。在我们之前的菌株中，使用最多20 g/L底物时可实现高产率（约为理论值的75%）乙醇生产。在本菌株中，使用高达50 g/L底物时可实现高产率乙醇生产。此外，引入[具体基因名称]使最大滴度提高了14%。
