生物技术通报 ›› 2023, Vol. 39 ›› Issue (11): 205-216.doi: 10.13560/j.cnki.biotech.bull.1985.2023-0722
收稿日期:
2023-07-28
出版日期:
2023-11-26
发布日期:
2023-12-20
通讯作者:
汪涯,男,博士,副教授,研究方向:微生物生态和代谢调控;E-mail: wangya@jxstnu.edu.cn作者简介:
唐瑞琪,女,博士,讲师,研究方向:微生物代谢工程和合成生物学;E-mail: rq_tang@jxstnu.edu.cn
基金资助:
TANG Rui-qi1(), ZHAO Xin-qing2, ZHU Du1, WANG Ya1()
Received:
2023-07-28
Published:
2023-11-26
Online:
2023-12-20
摘要:
木质纤维素类生物质是前景广阔的化石原料替代品,其生物炼制可生产生物能源、生物基化学品和生物材料等多种产品,可降低碳排放,有助于实现“双碳”目标,因此受到越来越多的关注。然而,木质纤维素生物炼制需要经过预处理、微生物发酵和产物纯化等多个步骤,其中,预处理过程产生的多种化合物抑制微生物的细胞生长和发酵性能,是制约生物转化效率的瓶颈之一。大肠杆菌是木质纤维素生物炼制常用的宿主,被广泛应用于多种化合物的生产,研究其对木质纤维素水解液中抑制物的耐受性,对于提高木质纤维素生物炼制效率具有重要意义。本文首先介绍了木质纤维素的主要成分和基本结构,对木质纤维素的预处理方法以及预处理后水解液中的主要抑制物种类进行了简单阐述;随后,总结了木质纤维素水解液中几类主要抑制物呋喃类、羧酸类和酚类对大肠杆菌细胞的毒性,以及大肠杆菌对上述抑制物的胁迫响应机制和基于机制的菌株改造靶点;最后,综述了提高大肠杆菌对上述抑制物的胁迫耐受性的菌株改造策略,包括随机突变、实验室适应性进化和组学辅助的理性设计等,为利用代谢工程构建用于木质纤维素生物炼制的高效大肠杆菌菌株提供参考。
唐瑞琪, 赵心清, 朱笃, 汪涯. 大肠杆菌对木质纤维素水解液抑制物的胁迫耐受性[J]. 生物技术通报, 2023, 39(11): 205-216.
TANG Rui-qi, ZHAO Xin-qing, ZHU Du, WANG Ya. Stress Tolerance of Escherichia coli to Inhibitors in Lignocellulosic Hydrolysates[J]. Biotechnology Bulletin, 2023, 39(11): 205-216.
图1 木质纤维素水解液中常见的抑制物及其来源 HMF *表示5-羟甲基糠醛
Fig. 1 Common inhibitors in lignocellulosic hydrolysates and their sources HMF * stands for 5-hydroxylmethylfurfural
基因 Gene | 敲除/过表达 Deletion/Overexpression | 描述 Description | 抑制物 Inhibitor | 参考文献 Reference |
---|---|---|---|---|
pgi, encoding glucose-6-phosphate isomerase | Deletion | Shunt glucose to pentose phosphate pathway to increase NADPH production | Furfural, HMF | [ |
pntAB, encoding pyridine nucleotide transhydrogenase | Overexpression | Convert NADP+ to NADPH using PntAB to increase NADPH regeneration | Furfural, HMF | [ |
yqhD, encoding aldehyde reductase | Deletion | Delete NADPH-dependent YqhD to reduce NADPH consumption | Furfural, HMF | [ |
yqhC, encoding transcriptional activator | Deletion | Delete YqhC to downregulate yqhDexpression, reducing NADPH consumption | Furfural | [ |
fucO, encoding propanediol oxidoreductase | Overexpression | Convert furfural using NADH-dependent FucO to reduce NADPH consumption | Furfural | [ |
yghA, encoding oxidoreductase | Overexpression | Convert furfural using NADH-dependent YghA to reduce NADPH consumption | Furfural, HMF | [ |
pncB and nadE, encoding NAD salvage pathway enzymes | Overexpression | Increase NAD(P)H level through the nicotine amide salvage pathway | Furfural | [ |
Heterologous xylBand BaBAD, encoding benzyl alcohol dehydrogenases | Overexpression | Convert furfural using NADH-dependent XylB and BaBAD to reduce NADPH consumption | Furfural | [ |
thyA, encoding thymidylate synthase | Overexpression | Overexpress ThyA to increase dTMP level for DNA repair | Furfural | [ |
potE and puuP, encoding polyamine transporters | Overexpression | Increase cytoplasmic polyamine level to maintain DNA synthesis | Furfural | [ |
lpcA, encoding D-sedoheptulose-7-phosphate isomerase | Overexpression | Overexpress LpcA to increase formation of lipopolysaccharides and NADPH | Furfural | [ |
pssA, encoding phosphatidylserine synthase | Overexpression | Increase phosphatidylethanolamine content to increase membrane integrity | Furfural, HMF | [ |
mdtJI, encoding multidrug resistance efflux pump | Overexpression | Export furfural by efflux pump MdtJI | Furfural, HMF | [ |
groESL, encoding chaperonin | Overexpression | Maintain proper folding of proteins | Furfural | [ |
表1 提高大肠杆菌呋喃类抑制物耐受性的基因靶点
Table 1 Gene targets for improving the tolerance of E. coli to furan inhibitors
基因 Gene | 敲除/过表达 Deletion/Overexpression | 描述 Description | 抑制物 Inhibitor | 参考文献 Reference |
---|---|---|---|---|
pgi, encoding glucose-6-phosphate isomerase | Deletion | Shunt glucose to pentose phosphate pathway to increase NADPH production | Furfural, HMF | [ |
pntAB, encoding pyridine nucleotide transhydrogenase | Overexpression | Convert NADP+ to NADPH using PntAB to increase NADPH regeneration | Furfural, HMF | [ |
yqhD, encoding aldehyde reductase | Deletion | Delete NADPH-dependent YqhD to reduce NADPH consumption | Furfural, HMF | [ |
yqhC, encoding transcriptional activator | Deletion | Delete YqhC to downregulate yqhDexpression, reducing NADPH consumption | Furfural | [ |
fucO, encoding propanediol oxidoreductase | Overexpression | Convert furfural using NADH-dependent FucO to reduce NADPH consumption | Furfural | [ |
yghA, encoding oxidoreductase | Overexpression | Convert furfural using NADH-dependent YghA to reduce NADPH consumption | Furfural, HMF | [ |
pncB and nadE, encoding NAD salvage pathway enzymes | Overexpression | Increase NAD(P)H level through the nicotine amide salvage pathway | Furfural | [ |
Heterologous xylBand BaBAD, encoding benzyl alcohol dehydrogenases | Overexpression | Convert furfural using NADH-dependent XylB and BaBAD to reduce NADPH consumption | Furfural | [ |
thyA, encoding thymidylate synthase | Overexpression | Overexpress ThyA to increase dTMP level for DNA repair | Furfural | [ |
potE and puuP, encoding polyamine transporters | Overexpression | Increase cytoplasmic polyamine level to maintain DNA synthesis | Furfural | [ |
lpcA, encoding D-sedoheptulose-7-phosphate isomerase | Overexpression | Overexpress LpcA to increase formation of lipopolysaccharides and NADPH | Furfural | [ |
pssA, encoding phosphatidylserine synthase | Overexpression | Increase phosphatidylethanolamine content to increase membrane integrity | Furfural, HMF | [ |
mdtJI, encoding multidrug resistance efflux pump | Overexpression | Export furfural by efflux pump MdtJI | Furfural, HMF | [ |
groESL, encoding chaperonin | Overexpression | Maintain proper folding of proteins | Furfural | [ |
基因 Gene | 描述 Description | 胁迫 Stress | 参考文献 Reference |
---|---|---|---|
dsrA and hfq, encoding small noncoding RNA and chaperone | DsrA increases rpoS mRNA stability and activate RpoS translation, Hfq promotes DsrA annealing to the rpoS5' untranscribed region(UTR) | Low pH | [ |
Heterologous cfaS, encoding cyclopropane fatty acid synthase | Decrease membrane permeability and fluidity | Low pH | [ |
Heterologous cbpA, encoding chaperone | CbpA plays a role in protein and DNA repair | Acetate | [ |
gadE, encoding transcriptional activator | GadE activates acid resistance system | Low pH | [ |
hdeB, encoding periplasmic acid stress chaperone | HdeB prevents periplasmic proteins aggregation at low pH | Low pH | [ |
sodB and katE, encoding superoxide dismutase and catalase | SodB and KatE are ROS scavengers | Low pH | [ |
表2 提高大肠杆菌耐酸性的过表达靶点
Table 2 Overexpression targets for improving the tolerance of E. coli to acid
基因 Gene | 描述 Description | 胁迫 Stress | 参考文献 Reference |
---|---|---|---|
dsrA and hfq, encoding small noncoding RNA and chaperone | DsrA increases rpoS mRNA stability and activate RpoS translation, Hfq promotes DsrA annealing to the rpoS5' untranscribed region(UTR) | Low pH | [ |
Heterologous cfaS, encoding cyclopropane fatty acid synthase | Decrease membrane permeability and fluidity | Low pH | [ |
Heterologous cbpA, encoding chaperone | CbpA plays a role in protein and DNA repair | Acetate | [ |
gadE, encoding transcriptional activator | GadE activates acid resistance system | Low pH | [ |
hdeB, encoding periplasmic acid stress chaperone | HdeB prevents periplasmic proteins aggregation at low pH | Low pH | [ |
sodB and katE, encoding superoxide dismutase and catalase | SodB and KatE are ROS scavengers | Low pH | [ |
图3 提高大肠杆菌对木质纤维素水解液中抑制物耐受性的示意图 HPLC/GC表示高效液相色谱或气相色谱,LC-/GC-MS表示液相或气相色谱与质谱联用,NMR表示核磁共振,ALE和ML分别表示实验室适应性进化和机器学习
Fig. 3 Schematic diagram of the tolerance improvement of E. coli to inhibitors in lignocellulosic hydrolysates HPLC/GC indicate high-performance liquid chromatography/gas chromatography, LC-/GC-MS indicate liquid chromatography-/gas chromatography-mass spectrometer, NMR indicates nuclear magnetic resonance, ALE indicates adaptive laboratory evolution, ML indicates machine learning
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