Article Open Access

Fed-batch Self-regulated Fermentation of Glucose to Co-produce Glycerol and 1,3-propanediol by Recombinant Escherichia coli

Synthetic Biology and Engineering. 2023, 1(2), 10008; https://doi.org/10.35534/sbe.2023.10008
Guimin Liu †,    Cai Feng †,    Zhiwei Zhu    Yaqin Sun    Zhilong Xiu *   
School of Bioengineering, Dalian University of Technology, Linggong Road 2#, Dalian 116024, China
These authors made equal contributions to the study.
*
Authors to whom correspondence should be addressed.

Received: 31 Mar 2023    Accepted: 16 May 2023    Published: 22 May 2023   

Abstract

As important bio-chemicals, glycerol and 1,3-propanediol (1,3-PDO) have been widely used in numerous fields, e.g., polymers, cosmetics, foods, lubricants, medicines, and so on. Bio-based 1,3-PDO is generally produced from glycerol or glucose by natural or recombinant strains, during which organic acids are always co-produced. In this work, acetic acid was also co-produced when 1,3-PDO was obtained from glucose by a recombinant strain of E. coli MG1655. Usually, a base was added to adjust the fermentation pH, resulting in the accumulation of organic salts and difficulty in the next down streaming process. Herein, a novel strategy was developed to consume the produced acetic acid by cells self in fed-batch self-regulated fermentation. The recombinant E. coli cells produced 48.33 g/L glycerol and 61.27 g/L 1,3-PDO with a total mass yield of 45.6% and without any other byproducts at the end of 5 fed-batch fermentations. The initial buffer pH, glucose concentration, pulse feeding sugar amount, time for a single batch fermentation and reducing acid were investigated by a series of comparative experiments. This fed-batch self-regulated fermentation has potential for the co-production of 1,3-PDO and glycerol, and will highlight the subsequent modification of recombinant E. coli strain by synthetic biology.

References

1.
Liu H, Xu Y, Zheng Z, Liu D. 1,3-Propanediol and its copolymers: Research, development and industrialization.  Biotechnol. J. 2010, 5, 1137–1148. [Google Scholar]
2.
Kaur G, Srivastava AK, Chand S. Advances in biotechnological production of 1,3-propanediol.  Biochem. Eng. J. 2012, 64, 106–118. [Google Scholar]
3.
Lee CS, Aroua MK, Daud WMAW, Cognet P, Pérès-Lucchese Y, Fabre P, et al. A review: Conversion of bioglycerol into 1,3-propanediol via biological and chemical method. Renew. Sustain. Energy Rev. 2015, 42, 963–972. [Google Scholar]
4.
Saxena RK, Anand P, Saran S, Isar J. Microbial production of 1,3-propanediol: Recent developments and emerging opportunities.  Biotechnol. Adv. 2009, 27, 895–913. [Google Scholar]
5.
Li X, Zhou Z, Li W, Yan Y, Shen X, Wang J, et al. Design of stable and self-regulated microbial consortia for chemical synthesis.  Nat. Commun. 2022, 13, 1554. [Google Scholar]
6.
Liu H, Zhang D, Xu Y, Mu Y, Sun Y, Xiu Z. Microbial production of 1,3-propanediol from glycerol by Klebsiella pneumoniae under micro-aerobic conditions up to a pilot scale.  Biotechnol. Lett. 2007, 29, 1281–1285. [Google Scholar]
7.
Sun YQ, Qi WT, Teng H, Xiu ZL, Zeng AP. Mathematical modeling of glycerol fermentation by Klebsiella pneumoniae: Concerning enzyme-catalytic reductive pathway and transport of glycerol and 1,3-propanediol across cell membrane.  Biochem. Eng. J. 2008, 38, 22–32.. [Google Scholar]
8.
Wilkens E, Ringel AK, Hortig D, Willke T, Vorlop K. High-level production of 1,3-propanediol from crude glycerol by Clostridium butyricum AKR102a.  Appl. Microbiol. Biotechnol. 2012, 93, 1057–1063. [Google Scholar]
9.
Zhou J, Shen J, Jiang L, Sun Y, Mu Y, Xiu Z. Selection and characterization of an anaerobic microbial consortium with high adaptation to crude glycerol for 1,3-propanediol production.  Appl. Microbiol. Biotechnol. 2017, 101, 5985–5996. [Google Scholar]
10.
Oh B, Lee S, Heo S, Seo J, Kim CH. Efficient production of 1,3-propanediol from crude glycerol by repeated fed-batch fermentation strategy of a lactate and 2,3-butanediol deficient mutant of Klebsiella pneumoniae Microb. Cell Fact. 2018, 17, 92. [Google Scholar]
11.
Wang X, Zhang L, Liang S, Yin Y, Wang P, Li Y, et al. Enhancing the capability of Klebsiella pneumoniae to produce 1,3-propanediol by overexpression and regulation through CRISPR‐dCas9.  Microb. Biotechnol. 2022, 15, 2112–2125. [Google Scholar]
12.
Wang W, Yu X, Wei Y, Ledesma-Amaro R, Ji X. Reprogramming the metabolism of Klebsiella pneumoniae for efficient 1,3-propanediol production.  Chem. Eng. Sci. 2021, 236, 116539. [Google Scholar]
13.
Sibley M, Ward JM. A cell engineering approach to enzyme-based fed-batch fermentation.  Microb. Cell Fact. 2021, 20, 146. [Google Scholar]
14.
Kuwae S, Ohda T, Tamashima H, Miki H, Kobayashi K. Development of a fed-batch culture process for enhanced production of recombinant human antithrombin by Chinese hamster ovary cells.  J. Biosci. Bioeng. 2005, 100, 502–510. [Google Scholar]
15.
Le H, Kabbur S, Pollastrini L, Sun Z, Mills K, Johnson K, et al. Multivariate analysis of cell culture bioprocess data—Lactate consumption as process indicator.  J. Biotechnol. 2012, 162, 210–223. [Google Scholar]
16.
Altamirano C, Paredes C, Illanes A, Cairó JJ, Gòdia F. Strategies for fed-batch cultivation of t-PA producing CHO cells: substitution of glucose and glutamine and rational design of culture medium.  J. Biotechnol. 2004, 110, 171–179. [Google Scholar]
17.
Li Y, Lin Z, Huang C, Zhang Y, Wang Z, Tang Y, et al. Metabolic engineering of Escherichia coli using CRISPR–Cas9 meditated genome editing.  Metab. Eng. 2015, 31, 13–21. [Google Scholar]
18.
Guo J, Wang T, Guan C, Liu B, Luo C, Xie Z, et al. Improved sgRNA design in bacteria via genome-wide activity profiling.  Nucleic Acids Res. 2018, 46, 7052–7069. [Google Scholar]
19.
Zhu Z, Zhou YJ, Kang MK, Krivoruchko A, Buijs NA, Nielsen J. Enabling the synthesis of medium chain alkanes and 1-alkenes in yeast.  Metab. Eng. 2017, 44, 81–88. [Google Scholar]
20.
Gibson DG, Young L, Chuang R, Venter JC, Hutchison CA, Smith HO. Enzymatic assembly of DNA molecules up to several hundred kilobases.  Nat. Methods 2009, 6, 343–345. [Google Scholar]
21.
Xia Y, Li K, Li J, Wang T, Gu L, Xun L. T5 exonuclease-dependent assembly offers a low-cost method for efficient cloning and site-directed mutagenesis.  Nucleic Acids Res. 2019, 47, 15. [Google Scholar]
22.
Heckman KL, Pease LR. Gene splicing and mutagenesis by PCR-driven overlap extension.  Nat. Protoc. 2007, 2, 924–932. [Google Scholar]
23.
Nakamura CE, Gatenby AA, Hsu AK, Reau RD, Haynie SL. Method for the production of 1,3-propanediol by recombinant microorganisms. US Patent 6,013,494, 2000.
24.
Wang X, Zhou J, Shen J, Zheng Y, Sun Y, Xiu Z. Sequential fed-batch fermentation of 1,3-propanediol from glycerol by Clostridium butyricum DL07.  Appl. Microbiol. Biotechnol. 2020, 104, 9179–9191. [Google Scholar]
25.
Zhou Y, Lu Z, Wang X, Selvaraj JN, Zhang G. Genetic engineering modification and fermentation optimization for extracellular production of recombinant proteins using Escherichia coli.  Appl. Microbiol. Biotechnol. 2018, 102, 1545–1556. [Google Scholar]
26.
Booth IR, Kroll RG. Regulation of cytoplasmic pH (pH1) in bacteria and its relationship to metabolism.  Biochem. Soc. Trans. 1983, 11, 70–72. [Google Scholar]
27.
Juhász T, Szengyel Z, Réczey K, Siika-Aho M, Viikari L. Characterization of cellulases and hemicellulases produced by Trichoderma reesei on various carbon sources.  Process Biochem. 2005, 40, 3519–3525. [Google Scholar]
28.
Murphy PT, Moore KJ, Richard TL, Bern CJ. Enzyme enhanced solid-state fermentation of kenaf core fiber for storage and pretreatment.  Bioresource Technol. 2007, 98, 3106–3111. [Google Scholar]
29.
Li C, He J, Chen H, Wang H, Han X, Wu Q. Optimization of fermentation conditions for producing neutral cellulase from a high-yield Bacillus megaterium genetic engineering bacteria.  J. Agric. Biotechnol. 2011, 19, 557–564. [Google Scholar]
30.
Liste-Calleja L, Lecina M, Lopez-Repullo J, Albiol J, Solà C, Cairó JJ. Lactate and glucose concomitant consumption as a self-regulated pH detoxification mechanism in HEK293 cell cultures.  Appl. Microbiol. Biotechnol. 2015, 99, 9951–9960. [Google Scholar]
31.
Li Z, Wu Z, Cen X, Liu Y, Zhang Y, Liu D, et al. Efficient Production of 1,3-Propanediol from Diverse Carbohydrates via a Non-natural Pathway Using 3-Hydroxypropionic Acid as an Intermediate.  ACS Synth. Biol. 2021, 10, 478–486. [Google Scholar]
32.
Chen J, Li W, Zhang Z, Tan T, Li Z. Metabolic engineering of Escherichia coli for the synthesis of polyhydroxyalkanoates using acetate as a main carbon source.  Microb. Cell Fact. 2018, 17, 102. [Google Scholar]
33.
Yang H, Zhang C, Lai N, Huang B, Fei P, Ding D, et al. Efficient isopropanol biosynthesis by engineered Escherichia coli using biologically produced acetate from syngas fermentation.  Bioresource Technol. 2020, 296, 122337. [Google Scholar]
34.
Li W, Chen J, Liu C, Yuan Q, Li Z. Microbial production of glycolate from acetate by metabolically engineered Escherichia coli J. Biotechnol. 2019, 291, 41–45. [Google Scholar]
35.
Lin H, Castro NM, Bennett GN, San K. Acetyl-CoA synthetase overexpression in Escherichia coli demonstrates more efficient acetate assimilation and lower acetate accumulation: a potential tool in metabolic engineering.  Appl. Microbiol. Biotechnol. 2006, 71, 870–874. [Google Scholar]
36.
Noh MH, Lim HG, Woo SH, Song J, Jung GY. Production of itaconic acid from acetate by engineering acid-tolerant Escherichia coli. W.  Biotechnol. Bioeng. 2018, 115, 729–738. [Google Scholar]
37.
Peng L, Shimizu K. Effect of fadR gene knockout on the metabolism of Escherichia coli based on analyses of protein expressions, enzyme activities and intracellular metabolite concentrations.  Enzyme Microb. Technol. 2006, 38, 512–520. [Google Scholar]
38.
Sato R, Tanaka T, Ohara H, Aso Y. Disruption of glpF gene encoding the glycerol facilitator improves 1,3-propanediol production from glucose via glycerol in Escherichia coli Lett. Appl. Microbiol. 2021, 72, 68–73. [Google Scholar]
39.
Szymanowska-Powalowska D, Bialas W. Scale-up of anaerobic 1,3-propanediol production by Clostridium butyricum DSP1 from crude glycerol.  BMC Microbiol. 2014, 14, 45. [Google Scholar]
40.
Wilkens E, Ringel AK, Hortig D, Willke T, Vorlop K. High-level production of 1,3-propanediol from crude glycerol by Clostridium butyricum AKR102a.  Appl. Microbiol. Biotechnol. 2012, 93, 1057–1063. [Google Scholar]
41.
Xue X, Li W, Li Z, Xia Y, Ye Q. Enhanced 1,3-propanediol production by supply of organic acids and repeated fed-batch culture.  J. Ind. Microbiol. Biot. 2010, 37, 681–687. [Google Scholar]
42.
Pflügl S, Marx H, Mattanovich D, Sauer M. Heading for an economic industrial upgrading of crude glycerol from biodiesel production to 1,3-propanediol by Lactobacillus diolivorans Bioresource Technol. 2014, 152, 499–504. [Google Scholar]
43.
Maina S, Kachrimanidou V, Ladakis D, Papanikolaou S, de Castro AM, Koutinas A. Evaluation of 1,3-propanediol production by two Citrobacter freundii strains using crude glycerol and soybean cake hydrolysate.  Environ. Sci. Pollut. Res. 2019, 26, 35523–35532. [Google Scholar]
44.
Zhou J, Shen J, Jiang L, Sun Y, Mu Y, Xiu Z. Selection and characterization of an anaerobic microbial consortium with high adaptation to crude glycerol for 1,3-propanediol production.  Appl. Microbiol. Biotechnol. 2017, 101, 5985–5996. [Google Scholar]
45.
Jiang L, Liu H, Mu Y, Sun Y, Xiu Z. High tolerance to glycerol and high production of 1,3-propanediol in batch fermentations by microbial consortium from marine sludge.  Eng. Life Sci. 2017, 17, 635–644. [Google Scholar]
46.
Wang X, Zhou J, Sun Y, Xiu Z. Bioconversion of Raw Glycerol from Waste Cooking-Oil-Based Biodiesel Production to 1,3-Propanediol and Lactate by a Microbial Consortium.  Front. Bioeng. Biotechnol. 2019, 7, 14. [Google Scholar]
47.
Liang Q, Zhang H, Li S, Qi Q. Construction of stress-induced metabolic pathway from glucose to 1,3-propanediol in Escherichia coli Appl. Microbiol. Biotechnol. 2011, 89, 57–62. [Google Scholar]
48.
Zhang Y, Li Z, Liu Y, Cen X, Liu D, Chen Z.  Systems metabolic engineering of Vibrio natriegens for the production of 1,3-propanediol.  Metab. Eng. 2021, 65, 52–65. [Google Scholar]
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