Spatial modulation and cofactor engineering of key pathway enzymes for fumarate production in <i>Candida glabrata</i>

Chen, Xiulai; Li, Yang; Tong, Tian; Liu, Li

doi:10.1002/bit.26906

Cited by 24 publications

(7 citation statements)

References 40 publications

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“…Regulation of PEP synthetase A and glyceraldehyde‐3‐phosphate dehydrogenase via promoter engineering has balanced the level of precursors required in isoprenoid biosynthesis in E. coli (Juyoung Jung et al, 2016). Similarly, promoter engineering of ADP‐dependent PEP carboxykinase and NAD + ‐dependent formate dehydrogenase fine‐tuned the cofactor balance for fumarate production in Candida glabrata (X. L. Chen et al, 2019). Hence, promoter engineering could effectively balance the supply of PEP and E4P.…”

Section: Introductionmentioning

confidence: 99%

Enhancing tryptophan production by balancing precursors in Escherichia coli

Guo

Ding

Liu

et al. 2021

Biotech & Bioengineering

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Tryptophan, an essential aromatic amino acid, is widely used in animal feed, food additives, and pharmaceuticals. Although sustainable and environmentally friendly, microbial tryptophan production from renewable feedstocks is limited by low biosynthesis and transport rates. Here, an Escherichia coli strain capable of efficient tryptophan production was generated by improving and balancing the supply of precursors and by engineering membrane transporters. Tryptophan biosynthesis was increased by eliminating negative regulatory factors, blocking competing pathways, and preventing tryptophan degradation. Promoter engineering balanced the supply of the precursors erythrose-4-phosphate and phosphoenolpyruvate, as well as the availability of serine. Finally, the engineering of tryptophan transporters prevented feedback inhibition and growth toxicity. Fed-batch fermentation of the final strain (TRP12) in a 5 L bioreactor produced 52.1 g•L −1 of tryptophan, with a yield of 0.171 g•g −1 glucose and productivity of 1.45 g•L −1 •h −1 . The metabolic engineering strategy described here paves the way for high-performance microbial cell factories aimed at the production of tryptophan as well as other valuable chemicals.

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Section: Introductionmentioning

confidence: 99%

Enhancing tryptophan production by balancing precursors in Escherichia coli

Guo

Ding

Liu

et al. 2021

Biotech & Bioengineering

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“…The reductive TCA pathway has been successfully applied to the biosynthesis of various organic acids by virtue of its high theoretical yield advantages. − For example, even if there was no decarboxylation step in the glyoxylic acid pathway, the theoretical yield would be only 1 mol of succinic acid per mol of glucose, but the theoretical yield using the reductive TCA pathway would be 2 mol of succinic acid per mol of glucose . The reason for this is that there is a carbon fixation reaction in the reductive TCA pathway.…”

Section: Discussionmentioning

confidence: 99%

Engineering the Reductive TCA Pathway to Dynamically Regulate the Biosynthesis of Adipic Acid in Escherichia coli

Hao

Zhou

et al. 2021

ACS Synth. Biol.

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Adipic acid is a versatile aliphatic dicarboxylic acid. It is applied mainly in the polymerization of nylon-6,6, which accounts for 50.8% of the global consumption market of adipic acid. The microbial production of adipic acid avoids the usage of petroleum resources and the emission of harmful nitrogen oxides that are generated by traditional chemical synthetic approaches. However, in the fermentation process, the low theoretical yield and the usage of expensive inducers hinders the large-scale industrial production of adipic acid. To overcome these challenges, we established an oxygen-dependent dynamic regulation (ODDR) system to control the expression of key genes (sucD, pyc, mdh, and f rdABCD) that could be induced to enhance the metabolic flux of the reductive TCA pathway under anaerobic conditions. Coupling of the constitutively expressed adipic acid synthetic pathway not only avoids the use of inducers but also increases the theoretical yield by nearly 50%. After the gene combination and operon structure were optimized, the reaction catalyzed by f rdABCD was found to be the rate-limiting step. Further optimizing the relative expression levels of sucD, pyc, and f rdABCD improved the titer of adipic acid 41.62-fold compared to the control strain Mad1415, demonstrating the superior performance of our ODDR system.

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“…Metabolic engineering is conducted to rewire the complete noncyclic glyoxylate pathway for fumarate production. Recently, five metabolic engineering strategies have been developed to enhance production of fumarate: reconstructing synthetic pathway, such as the reductive TCA cycle [ 2 ], the oxidative TCA cycle [ 3 ], the noncyclic glyoxylate cycle [ 4 ], the urea cycle and the purine nucleotide cycle [ 27 , 28 ]; eliminating byproducts formation [ 27 ], such as lactate, acetate, formate, malate, and succinate; optimizing oxidation and reduction levels [ 5 ]; modifying glucose transport system [ 8 ]; regulating C 4 -dicarboxylate transporter [ 26 ]. These results indicated that fumarate production has been improved by metabolic engineering strategies.…”

Section: Discussionmentioning

confidence: 99%

“…Recently, fumarate is mainly produced through three major metabolic pathways, including the reductive TCA cycle [ 2 ], the oxidative TCA cycle [ 3 ], and the noncyclic glyoxylate cycle [ 4 ]. The maximum theoretical yield of fumarate is 2 mol/mol glucose in reductive TCA cycle, but its fumarate productivity is limited due to two reversible reactions catalyzed by malate dehydrogenase and fumarase [ 5 ]. Based on this reductive TCA cycle, fumarate productivity was increased to 0.30 g/L/h by combinatorially regulating the expression of phosphoenolpyruvate carboxykinase and formate dehydrogenase [ 5 ].…”

Section: Introductionmentioning

confidence: 99%

“…The maximum theoretical yield of fumarate is 2 mol/mol glucose in reductive TCA cycle, but its fumarate productivity is limited due to two reversible reactions catalyzed by malate dehydrogenase and fumarase [ 5 ]. Based on this reductive TCA cycle, fumarate productivity was increased to 0.30 g/L/h by combinatorially regulating the expression of phosphoenolpyruvate carboxykinase and formate dehydrogenase [ 5 ]. In addition, fumarate production via the oxidative TCA cycle provides a maximum theoretical yield of 1 mol/mol glucose due to its release of 2 CO 2 .…”

Section: Introductionmentioning

confidence: 99%

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Engineering the transmission efficiency of the noncyclic glyoxylate pathway for fumarate production in Escherichia coli

Chen

Liu

et al. 2020

Biotechnol Biofuels

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Background Fumarate is a multifunctional dicarboxylic acid in the tricarboxylic acid cycle, but microbial engineering for fumarate production is limited by the transmission efficiency of its biosynthetic pathway. Results Here, pathway engineering was used to construct the noncyclic glyoxylate pathway for fumarate production. To improve the transmission efficiency of intermediate metabolites, pathway optimization was conducted by fluctuating gene expression levels to identify potential bottlenecks and then remove them, resulting in a large increase in fumarate production from 8.7 to 16.2 g/L. To further enhance its transmission efficiency of targeted metabolites, transporter engineering was used by screening the C 4 -dicarboxylate transporters and then strengthening the capacity of fumarate export, leading to fumarate production up to 18.9 g/L. Finally, the engineered strain E. coli W3110△4-P (H) CAI (H) SC produced 22.4 g/L fumarate in a 5-L fed-batch bioreactor. Conclusions In this study, we offered rational metabolic engineering and flux optimization strategies for efficient production of fumarate. These strategies have great potential in developing efficient microbial cell factories for production of high-value added chemicals.

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Spatial modulation and cofactor engineering of key pathway enzymes for fumarate production in Candida glabrata

Cited by 24 publications

References 40 publications

Enhancing tryptophan production by balancing precursors in Escherichia coli

Enhancing tryptophan production by balancing precursors in Escherichia coli

Engineering the Reductive TCA Pathway to Dynamically Regulate the Biosynthesis of Adipic Acid in Escherichia coli

Engineering the transmission efficiency of the noncyclic glyoxylate pathway for fumarate production in Escherichia coli

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