Improved poly-γ-glutamic acid production in Bacillus amyloliquefaciens by modular pathway engineering

Feng, Jun; Gu, Yanyan; Quan, Yufen; Cao, Mingfeng; Gao, Weixia; Zhang, Wei; Wang, Shufang; Yang, Chao; Song, Chunyan

doi:10.1016/j.ymben.2015.09.011

Cited by 88 publications

(87 citation statements)

References 53 publications

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“…With the current γ-PGA purification protocol, the produced levan cannot be separated from the desirable γ-PGA. The γ-PGA purity increased from 83.2 to 91.5% after s acB was deleted [33]. with the deletion of sacA in this work, the 1Δ group strains mostly used sacB and RBAM_031820 for sucrose metabolism and more levan was produced.…”

Section: Resultsmentioning

confidence: 90%

“…The γ-PGA production in all the strains was higher than that of the NK-1 strain (17.5–31.5% higher than NK-1 strain depending on different strains). From our previous experience [33], we knew that the levan synthesized by levansucrase is one of the main byproducts in γ-PGA production. With the current γ-PGA purification protocol, the produced levan cannot be separated from the desirable γ-PGA.…”

Section: Resultsmentioning

confidence: 99%

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Construction of energy-conserving sucrose utilization pathways for improving poly-γ-glutamic acid production in Bacillus amyloliquefaciens

Feng

Quan

et al. 2017

Microb Cell Fact

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BackgroundSucrose is an naturally abundant and easily fermentable feedstock for various biochemical production processes. By now, several sucrose utilization pathways have been identified and characterized. Among them, the pathway consists of sucrose permease and sucrose phosphorylase is an energy-conserving sucrose utilization pathway because it consumes less ATP when comparing to other known pathways. Bacillus amyloliquefaciens NK-1 strain can use sucrose as the feedstock to produce poly-γ-glutamic acid (γ-PGA), a highly valuable biopolymer. The native sucrose utilization pathway in NK-1 strain consists of phosphoenolpyruvate-dependent phosphotransferase system and sucrose-6-P hydrolase and consumes more ATP than the energy-conserving sucrose utilization pathway.ResultsIn this study, the native sucrose utilization pathway in NK-1 was firstly deleted and generated the B. amyloliquefaciens 3Δ strain. Then four combination of heterologous energy-conserving sucrose utilization pathways were constructed and introduced into the 3Δ strain. Results demonstrated that the combination of cscB (encodes sucrose permease) from Escherichia coli and sucP (encodes sucrose phosphorylase) from Bifidobacterium adolescentis showed the highest sucrose metabolic efficiency. The corresponding mutant consumed 49.4% more sucrose and produced 38.5% more γ-PGA than the NK-1 strain under the same fermentation conditions.ConclusionsTo our best knowledge, this is the first report concerning the enhancement of the target product production by introducing the heterologous energy-conserving sucrose utilization pathways. Such a strategy can be easily extended to other microorganism hosts for reinforced biochemical production using sucrose as substrate.Electronic supplementary materialThe online version of this article (doi:10.1186/s12934-017-0712-y) contains supplementary material, which is available to authorized users.

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Section: Resultsmentioning

confidence: 90%

Section: Resultsmentioning

confidence: 99%

Construction of energy-conserving sucrose utilization pathways for improving poly-γ-glutamic acid production in Bacillus amyloliquefaciens

Feng

Quan

et al. 2017

Microb Cell Fact

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“…Therefore, developing an efficient γ‐PGA production technology has the broad market prospect. Recently, many strategies have been attempted to improve the γ‐PGA yield, including the elimination of byproduct synthetic pathways, overexpression of glutamic acid synthesis pathways, and deletion of γ‐PGA hydrolase genes (Feng et al, , ; Scoffone et al, ). As the conversion of glutamic acid to γ‐PGA consumes ATP, a sufficient supply of ATP is crucial for a high‐level γ‐PGA synthesis (Su et al, ; Tang et al, ; Zhang et al, ).…”

Section: Discussionmentioning

confidence: 99%

Enhanced production of poly‐γ‐glutamic acid by improving ATP supply in metabolically engineered Bacillus licheniformis

Cai

Chen

et al. 2018

Biotech & Bioengineering

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Poly-γ-glutamic acid (γ-PGA) is an important multifunctional biopolymer with various applications, for which adenosine triphosphate (ATP) supply plays a vital role in biosynthesis. In this study, the enhancement of γ-PGA production was attempted through various approaches of improving ATP supply in the engineered strains of Bacillus licheniformis. The first approach is to engineer respiration chain branches of B. licheniformis, elimination of cytochrome bd oxidase branch reduced the maintenance coefficient, leading to a 19.27% increase of γ-PGA yield. The second approach is to introduce Vitreoscilla hemoglobin (VHB) into recombinant B. licheniformis, led to a 13.32% increase of γ-PGA yield. In the third approach, the genes purB and adK in ATP-biosynthetic pathway were respectively overexpressed, with the AdK overexpressed strain increased γ-PGA yield by 14.69%. Our study also confirmed that the respiratory nitrate reductase, NarGHIJ, is responsible for the conversion of nitrate to nitrite, and assimilatory nitrate reductase NasBC is for conversion of nitrite to ammonia. Both NarGHIJ and NasBC were positively regulated by the two-component system ResD-ResE, and overexpression of NarG, NasC, and ResD also improved the ATP supply and the consequent γ-PGA yield. Based on the above individual methods, a method of combining the deletion of cydBC gene and overexpression of genes vgB, adK, and resD were used to enhance ATP content of the cells to 3.53 μmol/g of DCW, the mutant WX-BCVAR with this enhancement produced 43.81 g/L of γ-PGA, a 38.64% improvement compared to wild-type strain WX-02. Collectively, our results demonstrate that improving ATP content in B. licheniformis is an efficient strategy to improve γ-PGA production.

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“…The cell concentration was monitored by measuring the absorbance at 600 nm which was then converted to Dry Cell Weight (DCW) by a calibration curve. The residual concentration of acetic acid in the culture was analyzed by HPLC (Feng et al, ). The remaining supernatants were filtered through a 0.22 µm filter and used for determination of residual glucose and amino acids, after appropriate dilution (the detection limitation for amino acids was 100 µM).…”

Section: Methodsmentioning

confidence: 99%

Metabolic engineering of Escherichia coli for microbial production of L‐methionine

Huang

Liu

Jin

et al. 2016

Biotech & Bioengineering

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L-methionine has attracted a great deal of attention for its nutritional, pharmaceutical, and clinical applications. In this study, Escherichia coli W3110 was engineered via deletion of a negative transcriptional regulator MetJ and over-expression of homoserine O-succinyltransferase MetA together with efflux transporter YjeH, resulting in L-methionine overproduction which is up to 413.16 mg/L. The partial inactivation of the L-methionine import system MetD via disruption of metI made the engineered E. coli ΔmetJ ΔmetI/pTrcA*H more tolerant to high L-ethionine concentration and accumulated L-methionine to a level 43.65% higher than that of E. coli W3110 ΔmetJ/pTrcA*H. Furthermore, deletion of lysA, which blocks the lysine biosynthesis pathway, led to a further 8.5-fold increase in L-methionine titer of E. coli ΔmetJ ΔmetI ΔlysA/pTrcA*H. Finally, addition of Na S O to the media led to an increase of fermentation titer of 11.45%. After optimization, constructed E. coli ΔmetJ ΔmetI ΔlysA/pTrcA*H was able to produce 9.75 g/L L-methionine with productivity of 0.20 g/L/h in a 5 L bioreactor. This novel metabolically tailored strain of E. coli provides an efficient platform for microbial production of L-methionine. Biotechnol. Bioeng. 2017;114: 843-851. © 2016 Wiley Periodicals, Inc.

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Improved poly-γ-glutamic acid production in Bacillus amyloliquefaciens by modular pathway engineering

Cited by 88 publications

References 53 publications

Construction of energy-conserving sucrose utilization pathways for improving poly-γ-glutamic acid production in Bacillus amyloliquefaciens

Construction of energy-conserving sucrose utilization pathways for improving poly-γ-glutamic acid production in Bacillus amyloliquefaciens

Enhanced production of poly‐γ‐glutamic acid by improving ATP supply in metabolically engineered Bacillus licheniformis

Metabolic engineering of Escherichia coli for microbial production of L‐methionine

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