Synthetic Biology Journal ›› 2023, Vol. 4 ›› Issue (6): 1161-1177.DOI: 10.12211/2096-8280.2023-051
• Invited Review • Previous Articles Next Articles
Huili SUN1,2,3, Jinyu CUI1,2,3, Guodong LUAN1,2,3, Xuefeng LYU1,2,3
Received:
2023-07-17
Revised:
2023-08-24
Online:
2024-01-19
Published:
2023-12-31
Contact:
Xuefeng LYU
孙绘梨1,2,3, 崔金玉1,2,3, 栾国栋1,2,3, 吕雪峰1,2,3
通讯作者:
吕雪峰
作者简介:
基金资助:
CLC Number:
Huili SUN, Jinyu CUI, Guodong LUAN, Xuefeng LYU. Progress of cyanobacterial synthetic biotechnology for efficient light-driven carbon fixation and ethanol production[J]. Synthetic Biology Journal, 2023, 4(6): 1161-1177.
孙绘梨, 崔金玉, 栾国栋, 吕雪峰. 面向高效光驱固碳产醇的蓝细菌合成生物技术研究进展[J]. 合成生物学, 2023, 4(6): 1161-1177.
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URL: https://synbioj.cip.com.cn/EN/10.12211/2096-8280.2023-051
策略 | 菌株 | 操作 | 效果 | 参考文献 |
---|---|---|---|---|
优化Adh2的活性 | PCC 6803 | 采用集胞藻PCC 6803来源的偏好使用NADPH为辅因子的slr1192基因替代运动发酵单胞菌adh2基因 | 增强了乙醛还原催化步骤与蓝细菌代谢背景的适配性,将乙醇产率提高了50% | [ |
优化Pdc与Adh2的实际丰度和配比 | PCC 6803 | 使用5种强度不同的RBS分别控制pdc与adh2的表达,并进行组合,评价其乙醇产量及酶活 | 发现乙醇合成通量更强烈地决定于pdc而非adh2的表达强度和丰度,在5种RBS控制的不同pdc表达强度的突变体中,pdc表达强度最高的藻株乙醇合成能力最强,培养7 d时的乙醇含量约为1 g/L | [ |
强化卡尔文循环以提高蓝细菌固碳效率并加强乙醇合成 | PCC 6803 | 分别将RuBisCo、SBPase、FBA或TK基因与Pdc-Adh2途径共表达 | 分别将乙醇产量提高了55%、67%、37%和69%,同时,总生物量也分别增加7.7%、15.1%、8.8%和10.1% | [ |
在表达Pdc-Adh2途径的基础上,共表达FBA与TK基因 | 相比于单独表达FBA藻株的乙醇产量提高9倍以上 | [ | ||
在表达Pdc-Adh2途径的基础上,共表达SBPase与FBA基因 | 相比FBA单独表达藻株的乙醇产量提高2.5倍 | [ | ||
加强蓝细菌底盘藻株对无机碳源的吸收 | PCC 7942 | 在表达Pdc-Adh2途径的基础上,过量表达ictB、ecaA以及groESL | 提高了在模拟烟道气条件下藻株的生物质含量(提高约4倍,0.9 g/L,72 h)和乙醇含量(提高约20倍,0.2 g/L,72 h) | [ |
敲除丙酮酸消耗途径,并将TCA循环中的碳流重新导回丙酮酸节点以增强乙醇合成前体供应 | PCC 6803 | 在表达Pdc-Adh2途径的基础上,敲除了PEP合成酶(PpsA)基因,催化糖原合成的关键酶(GlgC)基因,并过量表达大肠杆菌来源的苹果酸酶(MaeB)基因 | 乙醇产量提升至1.09 g/L(7 d) | [ |
阻断糖原合成途径 | PCC 7002 | 将两个乙醇合成途径(Pdc-Adh2)拷贝整合至基因组中两个糖原合成酶(GlgA)编码基因的位点,从而同时加强乙醇合成通量并阻断糖原合成途径竞争 | 在实验室环境柱式反应器中乙醇产量达到2.2 g/L(10 d),在户外挂袋式培养中乙醇产量达到0.8 g/L(7 d) | [ |
阻断糖原和PHB的合成途径 | PCC 6803 | 敲除糖原合成关键基因glgC | 乙醇产量从0.212 g/L(3 d)提高至0.297 g/L(3 d) | [ |
在以上基础上进一步敲除phaCE基因以阻断PHB合成路径 | 乙醇产量提高至0.332 g/L(3 d) | [ | ||
对以上藻株进行缺氮处理 | 乙醇产量达到0.6 g/L(3 d) | [ | ||
共培养“碳汇”工程策略 | PCC 6803 | 将敲除glgC和phaA基因以阻断糖原和PHB合成途径的藻株和整合了Pdc-Adh2途径的工程藻株进行共培养 | 双菌体系中的乙醇产量达到4.6 g/L(25 d),而单平台藻株(基因组同时进行乙醇合成途径整合和糖原、PHB合成途径阻断)的产量4.1 g/L(25 d) | [ |
补充还原力供应 | PCC 6803 | 过量表达内源G6PDH编码基因zwf,并导入Pdc-Adh2途径 | 乙醇产量从0.44 g/L增加到0.59 g/L(14 d),同时生物量积累增加了50% | [ |
向蓝细菌培养体系中添加金属氧化物以介导NADPH再生 | PCC 6803 | 在培养体系中添加MgO或Fe2O3 | 乙醇产量达到5.1 g/L或4.851 g/L(25 d) | [ |
区室化合成乙醇并靶向性模拟缺氮环境 | PCC 7120 | 在异形胞中使用特异性启动的hupS启动子控制Pdc-Adh2的表达 | 乙醇产量达到1.68 g/L(23 d) | [ |
在以上基础上,使用特异性靶向异形胞的CRISRPi基因表达系统抑制glnA的表达 | 乙醇产量提高了27% | [ | ||
通过优化的基因组尺度代谢网络模型,预测乙醇/生物量耦合突变体 | PCC 6803 | 预测最优突变体为通过13个基因的敲除来达到耦联乙醇合成和细胞生长的效果 | 预测乙醇产量为3.498 g/L(4 d) | [ |
Table 1 Optimization strategies for ethanol production by cyanobacteria photosynthesis
策略 | 菌株 | 操作 | 效果 | 参考文献 |
---|---|---|---|---|
优化Adh2的活性 | PCC 6803 | 采用集胞藻PCC 6803来源的偏好使用NADPH为辅因子的slr1192基因替代运动发酵单胞菌adh2基因 | 增强了乙醛还原催化步骤与蓝细菌代谢背景的适配性,将乙醇产率提高了50% | [ |
优化Pdc与Adh2的实际丰度和配比 | PCC 6803 | 使用5种强度不同的RBS分别控制pdc与adh2的表达,并进行组合,评价其乙醇产量及酶活 | 发现乙醇合成通量更强烈地决定于pdc而非adh2的表达强度和丰度,在5种RBS控制的不同pdc表达强度的突变体中,pdc表达强度最高的藻株乙醇合成能力最强,培养7 d时的乙醇含量约为1 g/L | [ |
强化卡尔文循环以提高蓝细菌固碳效率并加强乙醇合成 | PCC 6803 | 分别将RuBisCo、SBPase、FBA或TK基因与Pdc-Adh2途径共表达 | 分别将乙醇产量提高了55%、67%、37%和69%,同时,总生物量也分别增加7.7%、15.1%、8.8%和10.1% | [ |
在表达Pdc-Adh2途径的基础上,共表达FBA与TK基因 | 相比于单独表达FBA藻株的乙醇产量提高9倍以上 | [ | ||
在表达Pdc-Adh2途径的基础上,共表达SBPase与FBA基因 | 相比FBA单独表达藻株的乙醇产量提高2.5倍 | [ | ||
加强蓝细菌底盘藻株对无机碳源的吸收 | PCC 7942 | 在表达Pdc-Adh2途径的基础上,过量表达ictB、ecaA以及groESL | 提高了在模拟烟道气条件下藻株的生物质含量(提高约4倍,0.9 g/L,72 h)和乙醇含量(提高约20倍,0.2 g/L,72 h) | [ |
敲除丙酮酸消耗途径,并将TCA循环中的碳流重新导回丙酮酸节点以增强乙醇合成前体供应 | PCC 6803 | 在表达Pdc-Adh2途径的基础上,敲除了PEP合成酶(PpsA)基因,催化糖原合成的关键酶(GlgC)基因,并过量表达大肠杆菌来源的苹果酸酶(MaeB)基因 | 乙醇产量提升至1.09 g/L(7 d) | [ |
阻断糖原合成途径 | PCC 7002 | 将两个乙醇合成途径(Pdc-Adh2)拷贝整合至基因组中两个糖原合成酶(GlgA)编码基因的位点,从而同时加强乙醇合成通量并阻断糖原合成途径竞争 | 在实验室环境柱式反应器中乙醇产量达到2.2 g/L(10 d),在户外挂袋式培养中乙醇产量达到0.8 g/L(7 d) | [ |
阻断糖原和PHB的合成途径 | PCC 6803 | 敲除糖原合成关键基因glgC | 乙醇产量从0.212 g/L(3 d)提高至0.297 g/L(3 d) | [ |
在以上基础上进一步敲除phaCE基因以阻断PHB合成路径 | 乙醇产量提高至0.332 g/L(3 d) | [ | ||
对以上藻株进行缺氮处理 | 乙醇产量达到0.6 g/L(3 d) | [ | ||
共培养“碳汇”工程策略 | PCC 6803 | 将敲除glgC和phaA基因以阻断糖原和PHB合成途径的藻株和整合了Pdc-Adh2途径的工程藻株进行共培养 | 双菌体系中的乙醇产量达到4.6 g/L(25 d),而单平台藻株(基因组同时进行乙醇合成途径整合和糖原、PHB合成途径阻断)的产量4.1 g/L(25 d) | [ |
补充还原力供应 | PCC 6803 | 过量表达内源G6PDH编码基因zwf,并导入Pdc-Adh2途径 | 乙醇产量从0.44 g/L增加到0.59 g/L(14 d),同时生物量积累增加了50% | [ |
向蓝细菌培养体系中添加金属氧化物以介导NADPH再生 | PCC 6803 | 在培养体系中添加MgO或Fe2O3 | 乙醇产量达到5.1 g/L或4.851 g/L(25 d) | [ |
区室化合成乙醇并靶向性模拟缺氮环境 | PCC 7120 | 在异形胞中使用特异性启动的hupS启动子控制Pdc-Adh2的表达 | 乙醇产量达到1.68 g/L(23 d) | [ |
在以上基础上,使用特异性靶向异形胞的CRISRPi基因表达系统抑制glnA的表达 | 乙醇产量提高了27% | [ | ||
通过优化的基因组尺度代谢网络模型,预测乙醇/生物量耦合突变体 | PCC 6803 | 预测最优突变体为通过13个基因的敲除来达到耦联乙醇合成和细胞生长的效果 | 预测乙醇产量为3.498 g/L(4 d) | [ |
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