Overexpression of uracil permease and nucleoside transporter from Bacillus amyloliquefaciens improves cytidine production in Escherichia coli

Ma, Ruoshuang; Fang, Haitian; Liu, Huiyan; Pan, Lin; Wang, Hongyan; Zhang, Heng

doi:10.1007/s10529-021-03103-3

Cited by 6 publications

(4 citation statements)

References 29 publications

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“…Transporter engineering is one of the common strategies employed in metabolic engineering to enhance the synthesis of compounds in engineered cells. , It has been reported that UraA (uracil/H+ symporter), encoded by the uraA gene, forms a dimer through its interaction with uracil, facilitating the absorption of external uracil by E. coli, and PelB as a signal peptide can enhance the effect of UraA. , Thus, UraA and its coexpression with PelB were overexpressed in strain F4, resulting in strain G1 and strain G2.…”

Section: Resultsmentioning

confidence: 99%

Systematic Engineering of Escherichia coli for Efficient Production of Pseudouridine from Glucose and Uracil

Zhang,

Wei,

Zhou

et al. 2024

ACS Synth. Biol.

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In this study, we proposed a biological approach to efficiently produce pseudouridine (Ψ) from glucose and uracil in vivo using engineered Escherichia coli. By screening host strains and core enzymes, E. coli MG1655 overexpressing Ψ monophosphate (ΨMP) glycosidase and ΨMP phosphatase was obtained, which displayed the highest Ψ concentration. Then, optimization of the RBS sequences, enhancement of ribose 5-phosphate supply in the cells, and overexpression of the membrane transport protein UraA were investigated. Finally, fed-batch fermentation of Ψ in a 5 L fermentor can reach 27.5 g/L with a yield of 89.2 mol % toward uracil and 25.6 mol % toward glucose within 48 h, both of which are the highest to date. In addition, the Ψ product with a high purity of 99.8% can be purified from the fermentation broth after crystallization. This work provides an efficient and environmentally friendly protocol for allowing for the possibility of Ψ bioproduction on an industrial scale.

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Section: Resultsmentioning

confidence: 99%

Systematic Engineering of Escherichia coli for Efficient Production of Pseudouridine from Glucose and Uracil

Zhang,

Wei,

Zhou

et al. 2024

ACS Synth. Biol.

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“…Excessive accumulation of nucleosides leads to metabolic burden on the cell and causes productmediated feedback inhibition. Previous studies illustrated that the NupG protein of E. coli can transport extracellular nucleosides into cells, and knockout of the nupG and nupC genes can increase the uridine and cytidine titres [11,28,29]. Meanwhile, overexpression of the nucleoside e ux transporters PbuE and NepI notably increased inosine secretion [30].…”

Section: Transporter Engineering For Guanosine Accumulationmentioning

confidence: 99%

Efficient production of guanosine in Escherichia coli by combinatorial metabolic engineering

Zhang,

Qin,

Hou

et al. 2023

Preprint

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Guanosine is a purine nucleoside that is widely used as a raw material for food additives and pharmaceutical products. However, the lack of strains with efficiently producing guanosine greatly limited industrial application. We attempted to efficiently produce guanosinein Escherichia coli using systematic metabolic engineering. First, we overexpressed the purine synthesis pathway from Bacillus subtilis and the prs gene, and deleted three genes involved in guanosinecatabolism to increase guanosine accumulation. Subsequently, we attenuated purA expression and eliminated feedback inhibition. Then, we modified the metabolic flux of the glycolysis and Entner-Doudoroff pathways and performed redox cofactors rebalancing. Finally, transporter engineering further increased the guanosine titre to 123.6 mg/L. After 72 h of the fed-batch fermentation in shake-flask, the guanosine titre achieved 145.2 mg/L. Our results reveal that the guanosinesynthesis pathway was successfully optimized by combinatorial metabolic engineering, which could be applicable to the efficient synthesis of other nucleoside products.

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“…Excessive accumulation of nucleosides leads to metabolic burden on the cell and causes product-mediated feedback inhibition. Previous studies illustrated that the NupG protein of E. coli can transport extracellular nucleosides into cells, and knockout of the nupG and nupC genes can increase the uridine and cytidine titres [ 11 , 27 , 28 ]. Meanwhile, overexpression of the nucleoside efflux transporters PbuE and NepI notably increased inosine secretion [ 29 ].…”

Section: Blocking the Degradation Pathwaymentioning

confidence: 99%

Efficient production of guanosine in Escherichia coli by combinatorial metabolic engineering

Zhang,

Qin,

Hou

et al. 2024

Microb Cell Fact

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Background Guanosine is a purine nucleoside that is widely used as a raw material for food additives and pharmaceutical products. Microbial fermentation is the main production method of guanosine. However, the guanosine-producing strains possess multiple metabolic pathway interactions and complex regulatory mechanisms. The lack of strains with efficiently producing-guanosine greatly limited industrial application. Results We attempted to efficiently produce guanosine in Escherichia coli using systematic metabolic engineering. First, we overexpressed the purine synthesis pathway from Bacillus subtilis and the prs gene, and deleted three genes involved in guanosine catabolism to increase guanosine accumulation. Subsequently, we attenuated purA expression and eliminated feedback and transcription dual inhibition. Then, we modified the metabolic flux of the glycolysis and Entner-Doudoroff (ED) pathways and performed redox cofactors rebalancing. Finally, transporter engineering and enhancing the guanosine synthesis pathway further increased the guanosine titre to 134.9 mg/L. After 72 h of the fed-batch fermentation in shake-flask, the guanosine titre achieved 289.8 mg/L. Conclusions Our results reveal that the guanosine synthesis pathway was successfully optimized by combinatorial metabolic engineering, which could be applicable to the efficient synthesis of other nucleoside products.

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Overexpression of uracil permease and nucleoside transporter from Bacillus amyloliquefaciens improves cytidine production in Escherichia coli

Cited by 6 publications

References 29 publications

Systematic Engineering of Escherichia coli for Efficient Production of Pseudouridine from Glucose and Uracil

Systematic Engineering of Escherichia coli for Efficient Production of Pseudouridine from Glucose and Uracil

Efficient production of guanosine in Escherichia coli by combinatorial metabolic engineering

Efficient production of guanosine in Escherichia coli by combinatorial metabolic engineering

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