This is a preprint, a preliminary version of a manuscript that has not completed peer review at a journal. Research Square does not conduct peer review prior to posting preprints. The posting of a preprint on this server should not be interpreted as an endorsement of its validity or suitability for dissemination as established information or for guiding clinical practice.
Research
Wanmeng Mu
Jiangnan University
Corresponding Author
License:
This work is licensed under a CC BY 4.0 License. Read Full License
Background: Guanosine 5′-diphosphate (GDP)-L-fucose is a vital nucleotide sugar involved in the synthesis of fucosylated oligosaccharides, such as fucosylated human milk oligosaccharides, which play important roles in physiological and pathological processes.
Results: In this study, a combinatorial modular pathway engineering strategy was implemented to efficiently increase the intracellular titers of GDP-L-fucose in engineered Escherichia coli. The de novo GDP-L-fucose synthesis pathway was partitioned into two modules and fine-tuned in both transcriptional and translational levels, which remarkably improved the GDP-L-fucose production. In addition, the gene encoding the UDP-glucose lipid carrier transferase (WcaJ) was inactivated to eliminate the competing metabolite pathway from GDP-L-fucose to colanic acid. Furthermore, cofactors regeneration was underpinned to promote biocatalysis. Taken together, the final engineered strain EWL37, which could achieve the titers of 18.33 mg/L in shake-flask cultivation, was able to produce 106.21 mg/L (4.28 mg/g DCW) GDP-L-fucose through fed-batch cultivation.
Conclusion: To date, this is the first utilization of a modular pathway optimization approach to increase the production potential of the de novo synthesized GDP-L-fucose. In general, this study manifests that via combinatorial modular pathway engineering, the GDP-L-fucose biosynthesis can be significantly improved.
Keywords
GDP-L-fucose, modular pathway engineering, cofactor regeneration, the de novo pathway
Figure 1
Figure 2
Figure 3
Figure 4
Figure 5
Figure 6
This is a list of supplementary files associated with this preprint. Click to download.
Loading...
Badges
Prescreen
This manuscript passed our Prescreen assessment
Complete author information
Appropriate declaration statements
Potential risks to human health
Prescreen
History
CURRENT STATUS: Posted
Version 1
Posted 14 Jan, 2020
No community comments so far
Metrics
Comments: 0
PDF Downloads: ...
HTML Views: ...
Subject Areas
Biotechnology and Bioengineering
Learn more about our company.
This is a preprint, a preliminary version of a manuscript that has not completed peer review at a journal. Research Square does not conduct peer review prior to posting preprints. The posting of a preprint on this server should not be interpreted as an endorsement of its validity or suitability for dissemination as established information or for guiding clinical practice.
Research
Wanmeng Mu
Jiangnan University
Corresponding Author
Badges
Prescreen
This manuscript passed our Prescreen assessment
Complete author information
Appropriate declaration statements
Potential risks to human health
Prescreen
History
CURRENT STATUS: Posted
Version 1
Posted 14 Jan, 2020
No community comments so far
Metrics
Comments: 0
PDF Downloads: ...
HTML Views: ...
Subject Areas
Biotechnology and Bioengineering
License:
This work is licensed under a CC BY 4.0 License. Read Full License
Background: Guanosine 5′-diphosphate (GDP)-L-fucose is a vital nucleotide sugar involved in the synthesis of fucosylated oligosaccharides, such as fucosylated human milk oligosaccharides, which play important roles in physiological and pathological processes.
Results: In this study, a combinatorial modular pathway engineering strategy was implemented to efficiently increase the intracellular titers of GDP-L-fucose in engineered Escherichia coli. The de novo GDP-L-fucose synthesis pathway was partitioned into two modules and fine-tuned in both transcriptional and translational levels, which remarkably improved the GDP-L-fucose production. In addition, the gene encoding the UDP-glucose lipid carrier transferase (WcaJ) was inactivated to eliminate the competing metabolite pathway from GDP-L-fucose to colanic acid. Furthermore, cofactors regeneration was underpinned to promote biocatalysis. Taken together, the final engineered strain EWL37, which could achieve the titers of 18.33 mg/L in shake-flask cultivation, was able to produce 106.21 mg/L (4.28 mg/g DCW) GDP-L-fucose through fed-batch cultivation.
Conclusion: To date, this is the first utilization of a modular pathway optimization approach to increase the production potential of the de novo synthesized GDP-L-fucose. In general, this study manifests that via combinatorial modular pathway engineering, the GDP-L-fucose biosynthesis can be significantly improved.
Figure 1
Figure 2
Figure 3
Figure 4
Figure 5
Figure 6
This is a list of supplementary files associated with this preprint. Click to download.
Loading...