High-yield production of 1,3-propanediol from glycerol by metabolically engineered Klebsiella pneumoniae

Lee, Jung Hun; Jung, Moo Young; Oh, Min Kyu

doi:10.1186/s13068-018-1100-5

Cited by 56 publications

(14 citation statements)

References 39 publications

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“…There are many successful examples in the literature reporting cosubstrate fermentation to achieve better fermentation performance and to increase product yields. Adding sugars, mannitol, or hemicellulose hydrolysate as a cosubstrate to glycerol can significantly increase the yield of 1,3-propanediol production (31)(32)(33). Beet molasses added as a cosubstrate promoted 2,3-butanediol production from biodiesel derived glycerol (34).…”

Section: Discussionmentioning

confidence: 99%

Homoethanol Production from Glycerol and Gluconate Using Recombinant Klebsiella oxytoca Strains

Tao

Wang

Walters

et al. 2019

Appl Environ Microbiol

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Gluconic acid, an oxidized cellulose degradation product, could be produced from cellulosic biomass. Glycerol is an inexpensive and renewable resource for fuels and chemicals production and is available as a byproduct of biodiesel production. Gluconate is a more oxidized substrate than glucose, whereas glycerol is a more reduced substrate than glucose. Although the production of homoethanol from glucose can be achieved, the conversion of gluconate to ethanol is accompanied by the production of oxidized byproduct such as acetate, and reduced byproducts such as 1,3-propanediol are produced, along with ethanol, when glycerol is used as the carbon source. When gluconate and glycerol are used as the sole carbon source by Klebsiella oxytoca BW21, the ethanol yield is about 62 to 64%. Coutilization of both gluconate and glycerol in batch fermentation increased the yield of ethanol to about 78.7% and decreased by-product accumulation (such as acetate and 1,3-propanediol) substantially. Decreasing by-product formation by deleting the pta, frd, ldh, pflA, and pduC genes in strain BW21 increased the ethanol yield to 89.3% in the batch fermentation of a glycerol-gluconate mixture. These deletions produced the strain K. oxytoca WT26. However, the utilization rate of glycerol was significantly slower than that of gluconate in batch fermentation. In addition, substantial amounts of glycerol remain unutilized after gluconate was depleted in batch fermentation. Continuous fed-batch fermentation was used to solve the utilization rate mismatch problem for gluconate and glycerol. An ethanol yield of 97.2% was achieved in continuous fed-batch fermentation of these two substrates, and glycerol was completely used at the end of the fermentation. IMPORTANCE Gluconate is a biomass-derived degradation product, and glycerol can be obtained as a biodiesel byproduct. Compared to glucose, using them as the sole substrate is accompanied by the production of by-products. Our study shows that through pathway engineering and adoption of a fed-batch culture system, high-yield homoethanol production that usually can be achieved by using glucose as the substrate is achievable using gluconate and glycerol as cosubstrates. The same strategy is expected to be able to achieve homofermentative production of other products, such as lactate and 2,3-butanediol, which can be typically achieved using glucose as the substrate and inexpensive biodiesel-derived glycerol and biomass-derived gluconate as the cosubstrates.

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Section: Discussionmentioning

confidence: 99%

Homoethanol Production from Glycerol and Gluconate Using Recombinant Klebsiella oxytoca Strains

Tao

Wang

Walters

et al. 2019

Appl Environ Microbiol

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“…In this study, we have leveraged the protein arraying approach, used natively by several cellulolytic bacteria to tether their biomass degrading enzymes, to self-organize metabolic pathways (including sensitive enzymes) and simplify their purifications into a single step from a co-expressing enzyme production strain (Figure 1). We applied this approach to the production of 1,3-propanediol (1,3-PDO) from glycerol (Raynaud et al, 2003;Jiang et al, 2016;Lee et al, 2018). Glycerol, a main byproduct of the biofuel industry, represents an ideal conversion intermediate that can be upgraded to a valueadded product such as 1,3-PDO (Çelik et al, 2008;Yang et al, 2012;Plácido and Capareda, 2016).…”

Section: Introductionmentioning

confidence: 99%

Self-Assembling Metabolon Enables the Cell Free Conversion of Glycerol to 1,3-Propanediol

Alahuhta

Hewitt

et al. 2021

Front. Energy Res.

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Cell free biocatalysis is showing promise as a replacement or complement to conventional microbial biocatalysts due to the potential for achieving high yields, titers, and productivities. However, there exist several challenges that need to be addressed before its broader industrial adoption is achieved. New paradigms and innovative solutions are needed to overcome these challenges. In this study we demonstrate high levels of glycerol conversion to 1,3-propanediol using a self-assembling metabolic pathway leveraging the arraying strategy (protein scaffolds) used by thermophilic cellulolytic bacteria to assemble their biomass degrading enzymes. These synthetic metabolons were capable of producing 1,3-PDO at a yield more than 95% at lower glycerol concentration and close to 70% at higher concentrations at a higher productivity rate than the equivalent microbial strain. One of the benefits of our approach is the fact that no enzyme purification is required, and that the assembly of the complex is accomplished in vivo before immobilization, while product formation is conducted in vitro. We also report the recovery of enzymatic activity upon fusion enzymes binding to these protein scaffolds, which could have broader applications when assembling arrayed protein complexes.

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“…Nevertheless, considerable amounts of by-product metabolites, especially lactate, acetic acid and ethanol [9], are synthesized via an oxidative pathway with pyruvate as the precursor. Therefore, elimination of by-product formation is a major strategy to metabolically engineer microorganism to improve 1,3-PD yield [10]. Deletions of genes of ethanol dehydrogenase (adhE) for ethanol synthesis,acetate kinase (ack) for acetic acid synthesis or lactate dehydrogenase (ldhA) for lactate synthesis promoted 1,3-PD production in K. pneumonia [11].…”

Section: Introductionmentioning

confidence: 99%

Klebsiella Pneumoniae Metabolic Regulation and its Application in 1,3-Propanediol Production

Wang¹,

Mu²,

Liu³

et al. 2021

Preprint

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Klebsiella pneumoniae is a well-known model organism for glycerol metabolism to produce 1,3-propanediol (1,3-PD), a valuable chemical intermediate for materials, such as polyesters. However, the relatively low conversion rate and productivity, as well as the accumulation of by-products such as lactic acid, ethanol and acetic acid, inhibit the production of 1,3-PD. Hereby, the 1,3-PD metabolism in K. pneumoniae was regulated through pathway engineering by using CRISPR-Cas9 technology for the first time to knock out the ldhA gene of lactate dehydrogenase,the adhE gene of alcohol dehydrogenase and the ack gene of acetate kinase respectively as needed and constructed recombinant bacteria ldhA(−), ldhA(−)-ack(−), ldhA(−)-adhE(−) and ldhA(−)-adhE(−)-ack(−), all of which showed a decrease in by-product production, leading to a higher NADH availability, and 1,3-BD production was significantly increased. In the shake flask fermentation, the 1,3-PD yield and conversion rate of the recombinant strain ldhA(−), ldhA(−)-ack(−), ldhA(−)-adhE(−), ldhA(−)-adhE(−)-ack(−) were higher than those of the parent strain. In the fed-batch fermentation, the 1,3-PD yield and conversion rate of the recombinant strain ldhA(−) were higher than those of the parent strain. The biomass of the recombinant strain ldhA(−)-adhE(−)-ack(−) was reduced due to the accumulation of acetic acid, but its 1,3-PD conversion rate was still higher than that of the parent strain. The higher productivity and fewer by-products concluded that the four Klebsiella pneumoniae recombinant strains could be promising industrial strain for economical production of 1,3-PD.

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High-yield production of 1,3-propanediol from glycerol by metabolically engineered Klebsiella pneumoniae

Cited by 56 publications

References 39 publications

Homoethanol Production from Glycerol and Gluconate Using Recombinant Klebsiella oxytoca Strains

Homoethanol Production from Glycerol and Gluconate Using Recombinant Klebsiella oxytoca Strains

Self-Assembling Metabolon Enables the Cell Free Conversion of Glycerol to 1,3-Propanediol

Klebsiella Pneumoniae Metabolic Regulation and its Application in 1,3-Propanediol Production

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