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Engineering Escherichia coli for a high yield of 1,3-propanediol near the theoretical maximum through chromosomal integration and gene deletion

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Abstract

Glycerol dehydratase (gdrAB-dhaB123) operon from Klebsiella pneumoniae and NADPH-dependent 1,3-propanediol oxidoreductase (yqhD) from Escherichia coli were stably integrated on the chromosomal DNA of E. coli under the control of the native-host ldhA and pflB constitutive promoters, respectively. The developed E. coli NSK015 (∆ldhA::gdrAB-dhaB123ackA::FRT ∆pflB::yqhDfrdABCD::cat-sacB) produced 1,3-propanediol (1,3-PDO) at the level of 36.8 g/L with a yield of 0.99 mol/mol of glycerol consumed when glucose was used as a co-substrate with glycerol. Co-substrate of glycerol and cassava starch was also utilized for 1,3-PDO production with the concentration and yield of 31.9 g/L and 0.84 mol/mol of glycerol respectively. This represents a work for efficient 1,3-PDO production in which the overexpression of heterologous genes on the E. coli host genome devoid of plasmid expression systems. Plasmids, antibiotics, IPTG, and rich nutrients were omitted during 1,3-PDO production. This may allow a further application of E. coli NSK015 for the efficient 1,3-PDO production in an economically industrial scale.

Key points

 • gdrAB-dhaB123 and yqhD were overexpressed in E. coli devoid of a plasmid system

E. coli NSK015 produced a high yield of 1,3-PDO at 99% theoretical maximum

Cassava starch was alternatively used as substrate for economical 1,3-PDO production

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Data sharing not applicable to this article as no datasets were generated or analyzed during the current study.

References

  • Bradford MM (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72:248–254

    Article  CAS  PubMed  Google Scholar 

  • Chen Z, Liu H, Liu D (2011) Metabolic pathway analysis of 1,3-propanediol production with a genetically modified Klebsiella pneumoniae by overexpressing an endogenous NADPH-dependent alcohol dehydrogenase. Biochem Eng J 54:151–157

    Article  CAS  Google Scholar 

  • Cintolesi A, Clomburg JM, Rigou V, Zygourakis K, Gonzalez R (2012) Quantitative analysis of the fermentative metabolism of glycerol in Escherichia coli. Biotechnol Bioeng 109:187–198

    Article  CAS  PubMed  Google Scholar 

  • Conway T, Sewell GW, Osman YA, Ingram LO (1987) Cloning and sequencing of the alcohol dehydrogenase II gene from Zymomonas mobilis. J Bacteriol 169:2591–2597

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Datsenko KA, Wanner BL (2000) One-step inactivation of chromosomal genes in Escherichia coli K-12 using PCR products. Proc Natl Acad Sci USA 97:6640–6645

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Dharmadi Y, Murarka A, Gonzalez R (2006) Anaerobic fermentation of glycerol by Escherichia coli: a new platform for metabolic engineering. Biotechnol Bioeng 94:821–829

    Article  CAS  PubMed  Google Scholar 

  • Di Laccio E, Elling RA, Wilson DK (2006) Identification of a novel NADH-specific aldo-keto reductase using sequence and structural homologies. Biochem J 400:105–114

    Article  Google Scholar 

  • Emptage M, Haynie SL, Laffend LA, Pucci JP, Whited G (2003) Process for the biological production of 1,3-propanediol with high titer. United States Patent, US6514733B1

  • Hong E, Kim J, Ha S, Ryu Y (2015) Improved 1,3-propanediol production by Escherichia coli from glycerol due to Co-expression of glycerol dehydratase reactivation factors and succinate addition. Biotechnol Bioprocess Eng 20:849–855

    Article  CAS  Google Scholar 

  • In S, Khunnonkwao P, Wong N, Phosiran C, Jantama SS, Jantama K (2020) Combining metabolic engineering and evolutionary adaptation in Klebsiella oxytoca KMS004 to significantly improve optically pure D-(−)-lactic acid yield and specific productivity in low nutrient medium. Appl Microbiol Biotechnol 104:9565–9579

    Article  CAS  PubMed  Google Scholar 

  • Jampatesh S, Sawisit A, Wong N, Jantama SS, Jantama K (2019) Evaluation of inhibitory effect and feasible utilization of dilute acid-pretreated rice straws on succinate production by metabolically engineered Escherichia coli AS1600a. Bioresourc Technol 273:93–102

    Article  CAS  Google Scholar 

  • Jantama K, Zhang X, Moore JC, Shanmugam KT, Svoronos SA, Ingram LO (2008) Eliminating side products and increasing succinate yields in engineered strains of Escherichia coli C. Biotechnol Bioeng 101:881–893

    Article  CAS  PubMed  Google Scholar 

  • Jantama K, Polyiam P, Khunnonkwao P, Chan S, Sangproo M, Khor K, Jantama SS, Kanchanatawee S (2015) Efficient reduction of the formation of by-products and improvement of production yield of 2,3-butanediol by a combined deletion of alcohol dehydrogenase, acetate kinase-phosphotransacetylase, and lactate dehydrogenase genes in metabolically engineered Klebsiella oxytoca in mineral salts medium. Metab Eng 30:16–26

    Article  CAS  PubMed  Google Scholar 

  • Jiang W, Zhuang Y, Wang S, Fang B (2015) Directed evolution and resolution mechanism of 1, 3-propanediol oxidoreductase from Klebsiella pneumoniae toward higher activity by error-prone PCR and bioinformatics. PLoS ONE 10:1–10. https://doi.org/10.1371/journal.pone.0141837

    Article  CAS  Google Scholar 

  • Khor K, Sawisit A, Chan S, Kanchanatawee S, Jantama SS, Jantama K (2016) High production yield and specific productivity of succinate from cassava starch by metabolically engineered Escherichia coli KJ122. J Chem Technol Biotechnol 91:2834–2841

    Article  CAS  Google Scholar 

  • Khunnonkwao P, Jantama SS, Kanchanatawee S, Jantama K (2018) Re-engineering Escherichia coli KJ122 to enhance the utilization of xylose and xylose/glucose mixture for efficient succinate production in mineral salt medium. Appl Microbiol Biotechnol 102:127–141

    Article  CAS  PubMed  Google Scholar 

  • Khunnonkwao P, Jantama SS, Jantama K, Joannis-Cassan C, Taillandier P (2020) Sequential coupling of enzymatic hydrolysis and fermentation platform for high yield and economical production of 2,3-butanediol from cassava by metabolically engineered Klebsiella oxytoca. J Chem Technol Biotechnol 96:1292–1301

    Article  CAS  Google Scholar 

  • Kim K, Kim SK, Park YC, Seo JH (2014) Enhanced production of 3-hydroxypropionic acid from glycerol by modulation of glycerol metabolism in recombinant Escherichia coli. Bioresour Technol 156:170–175

    Article  CAS  PubMed  Google Scholar 

  • Knietsch A, Bowien S, Whited G, Gottschalk G, Daniell R (2003) Identification and characterization of coenzyme B12-dependent glycerol dehydratase- and diol dehydratase-encoding genes from metagenomic DNA libraries derived from enrichment cultures. Appl Environ Microbiol 69:3048–3060

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Koma D, Yamanaka H, Moriyoshi K, Ohmoto SK (2012) Production of aromatic compounds by metabolically engineered Escherichia coli with an expanded shikimate pathway. Appl Environ Microbiol 78:6203–6216

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Lee JH, Jung MY, Oh MK (2018a) High-yield production of 1,3-propanediol from glycerol by metabolically engineered Klebsiella pneumoniae. Biotechnol Biofuels 11:1–13

    Article  CAS  Google Scholar 

  • Lee JH, Lama S, Kim JR, Park SH (2018b) Production of 1,3-propanediol from glucose by recombinant Escherichia coli BL21(DE3). Biotechnol Bioprocess Eng 23:250–258

    Article  CAS  Google Scholar 

  • Li H, Chen J, Li Y (2008) Enhanced activity of yqhD oxidoreductase in synthesis of 1,3-propanediol by error-prone PCR. Prog Nat Sci 18:1519–1524

    Article  CAS  Google Scholar 

  • Liang Q, Zhang H, Li S, Qi Q (2011) Construction of stress-induced metabolic pathway from glucose to 1,3-propanediol in Escherichia coli. Appl Microbiol Biotechnol 89:57–62

    Article  CAS  PubMed  Google Scholar 

  • Lim HG, Noh MH, Jeong JH, Park S, Jung GY (2016) Optimum rebalancing of the 3-hydroxypropionic acid production pathway from glycerol in Escherichia coli. ACS Synth Biol 5:1247–1255

    Article  CAS  PubMed  Google Scholar 

  • Liu H, Xu Y, Zheng Z, Liu D (2010) 1,3-Propanediol and its copolymers: research, development and industrialization. Biotechnol J 5:1137–1148

    Article  CAS  PubMed  Google Scholar 

  • Ma Z, Rao Z, Xu L, Liao X, Fang H, Zhuge B, Zhuge J (2009) Production of 1,3-propanediol from glycerol by engineered Escherichia coli using a novel co-expression vector. African J Biotechnol 8:5489–5494

    CAS  Google Scholar 

  • Martinez A, Grabar TB, Shanmugam KT, Yomano LP, York SW, Ingram LO (2007) Low salt medium for lactate and ethanol production by recombinant Escherichia coli B. Biotechnol Lett 29:397–404

    Article  CAS  PubMed  Google Scholar 

  • McCloskey D, Xu J, Schrübbers L, Christensen HB, Herrgård MJ (2018) RapidRIP quantifies the intracellular metabolome of 7 industrial strains of E. coli. Metab Eng 47:383–392

    Article  CAS  PubMed  Google Scholar 

  • Oh BR, Lee SM, Heo SY, Seo JW, Kim CH (2018) 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 17:1–9. https://doi.org/10.1186/s12934-018-0921-z

    Article  CAS  Google Scholar 

  • Perez JM, Arenas FA, Pradenas GA, Sandoval JM, Vasquez CC (2008) Escherichia coli YqhD exhibits aldehyde reductase activity and protects from the harmful effect of lipid peroxidation-derived aldehydes. J Bio Chem 283:7346–7353

    Article  CAS  Google Scholar 

  • Przystałowska H, Zeyland J, Szymanowska-Powałowska D, Szalata M, Słomski R, Lipiński D (2015) 1,3-propanediol production by new recombinant Escherichia coli containing genes from pathogenic bacteria. Microbiol Res 171:1–7

    Article  PubMed  CAS  Google Scholar 

  • Rao Z, Ma Z, Shen W, Fang H, Zhuge J, Wang X (2008) Engineered Saccharomyces cerevisiae that produces 1,3-propanediol from D-glucose. J Appl Microbiol 105:1768–1776

    Article  CAS  PubMed  Google Scholar 

  • Rathnasingh C, Raj SM, Jo JE, Park S (2009) Development and evaluation of efficient recombinant Escherichia coli strains for the production of 3-hydroxypropionic acid from glycerol. Biotechnol Bioeng 104:729–739

    CAS  PubMed  Google Scholar 

  • Rodriguez G, Atsumi S (2012) Isobutyraldehyde production from Escherichia coli by removing aldehyde reductase activity. Microb Cell Fact 11:90

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Sawisit A, Jampatesh S, Jantama SS, Jantama K (2018) Optimization of sodium hydroxide pretreatment and enzyme loading for efficient hydrolysis of rice straw to improve succinate production by metabolically engineered Escherichia coli KJ122 under simultaneous saccharification and fermentation. Bioresourc Technol 260:348–356

    Article  CAS  Google Scholar 

  • Seo JW, Seo MY, Oh BR, Heo SY, Baek JO, Rairakhwada D, Luo LH, Hong WK, Kim CH (2010) Identification and utilization of a 1,3-propanediol oxidoreductase isozyme for production of 1,3-propanediol from glycerol in Klebsiella pneumoniae. Appl Microbiol Biotechnol 85:659–666

    Article  CAS  PubMed  Google Scholar 

  • Skraly FA, Lytle BL, Cameron DC (1998) Construction and characterization of a 1,3-propanediol operon. Appl Environ Microbiol 64:98–105

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Tang X, Tan Y, Zhu H, Zhao K, Shen W (2009) Microbial conversion of glycerol to 1,3-propanediol by an engineered strain of Escherichia coli. Appl Environ Microbiol 75:1628–1634

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Tong IT, Cameron DC (1992) Enhancement of 1,3-propanediol production by cofermentation in Escherichia coli expressing Klebsiella pneumoniae dha regulon genes. Appl Biochem Biotechnol 34:149–159

    Article  PubMed  Google Scholar 

  • Tong IT, Liao HH, Cameron DC (1991) 1,3-Propanediol production by Escherichia coli expressing genes from the Klebsiella pneumoniae dha regulon. Appl Environ Microbiol 57:3541–3546

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Wang F, Qu H, Zhang D, Tian P, Tan T (2007) Production of 1,3-propanediol from glycerol by recombinant Escherichia coli using incompatible plasmids system. Mol Biotechnol 37:112–119

    Article  CAS  PubMed  Google Scholar 

  • Yang B, Liang S, Liu H, Liu J, Cui Z, Wen J (2018) Metabolic engineering of Escherichia coli for 1,3-propanediol biosynthesis from glycerol. Bioresour Technol 267:599–607

    Article  CAS  PubMed  Google Scholar 

  • Yun J, Yang M, Magocha TA, Zhang H, Xue Y, Zhang G, Qi X, Sun W (2018) Production of 1,3-propanediol using a novel 1,3-propanediol dehydrogenase from isolated Clostridium butyricum and co-biotransformation of whole cells. Bioresour Technol 247:838–843

    Article  CAS  PubMed  Google Scholar 

  • Yun J, Zabed HM, Zhang Y, Parvez A, Zhang G, Qi X (2021) Co-fermentation of glycerol and glucose by a co-culture system of engineered Escherichia coli strains for 1,3-propanediol production without vitamin B12 supplementation. Bioresour Technol 319:124218

    Article  CAS  PubMed  Google Scholar 

  • Zhu MM, Lawman PD, Cameron DC (2002) Improving 1,3-propanediol production from glycerol in a metabolically engineered Escherichia coli by reducing accumulation of sn-glycerol-3-phosphate. Biotechnol Prog 18:694–699

    Article  CAS  PubMed  Google Scholar 

  • Zhu JG, Li S, Ji XJ, Huang H, Hu N (2009) Enhanced 1,3-propanediol production in recombinant Klebsiella pneumoniae carrying the gene ydhD encoding 1,3-propanediol oxidoreductase isozyme. World J Microbiol Biotechnol 25:1217–1223

    Article  CAS  Google Scholar 

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Acknowledgements

Author thanks the Thailand Research Fund (TRF) under the Royal Golden Jubilee PhD scholarship (Grant No. PHD/0125/2556) that provided a financial support for this work.

Funding

This study was funded by Thailand Research Fund (TRF) under the Royal Golden Jubilee PhD scholarship (Grant No. PHD/0125/2556).

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KJ conceived, designed research, and performed project administration and funding acquisition. NW conducted experiments and wrote an original draft of the manuscript. KJ also analyzed data, provided comments, and edited and reviewed the final manuscript. All authors read and approved the manuscript.

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Correspondence to Kaemwich Jantama.

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Wong, N., Jantama, K. Engineering Escherichia coli for a high yield of 1,3-propanediol near the theoretical maximum through chromosomal integration and gene deletion. Appl Microbiol Biotechnol 106, 2937–2951 (2022). https://doi.org/10.1007/s00253-022-11898-y

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