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Research
Weifeng Liu
Institute of Microbiology Chinese Academy of Sciences
Corresponding Author
ORCiD: https://orcid.org/0000-0001-7218-3932
License:
This work is licensed under a CC BY 4.0 License. Read Full License
Background
Phenylpropanoid including raspberry ketone, is a kind of important natural plant product and widely used in pharmaceuticals, chemicals, cosmetics, and healthcare products. Bioproduction of phenylpropanoid in Escherichia coli and other microbial cell factories is an attractable approach considering the low phenylpropanoid contents in plants. However, it is usually difficult to produce high titer phenylpropanoid production when fermentation using glucose as carbon source. Developing novel bioprocess using alternative sources might provide a solution to this problem. In this study, typical phenylpropanoid raspberry ketone was used as the target product to develop a biosynthesis pathway for phenylpropanoid production from fatty acids, a promising alternative low-cost feedstock.
Results
A raspberry ketone biosynthesis module was developed and optimized by introducing 4-coumarate-CoA ligase (4CL), benzalacetone synthase (BAS), and raspberry ketone reductase (RZS) in Escherichia coli strains CR1-CR4. Then strain CR5 was developed by introducing raspberry ketone biosynthesis module into a fatty acids-utilization chassis FA09 to achieve production of raspberry ketone from fatty acids feedstock. However, the production of raspberry ketone was still limited by the low biomass and unable to substantiate whole-cell bioconversion process. Thus, a process by coordinately using fatty-acids and glycerol was developed. In addition, we systematically screened and optimized fatty acids-response promoters. The optimized promoter Pfrd3 was then successfully used for the efficient expression of key enzymes of raspberry ketone biosynthesis module during bioconversion from fatty acids. The final engineered strain CR8 could efficiently produce raspberry ketone repeatedly using bioconversion from fatty acids feedstock strategy, and about 291.3 mg/L raspberry ketone was produced.
Conclusion
Metabolically engineered Escherichia coli strains were successfully developed for raspberry ketone production from fatty acids using several strategies, including optimization of bioconversion process and fine-tuning key enzyme expression. This study provides an essential reference to establish the low-cost biological manufacture of phenylpropanoids compounds.
Keywords
raspberry ketone, fatty acids feedstock, bioconversion, phenylpropanoids, Escherichia coli
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View the published article at Microbial Cell Factories.
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On 28 Dec, 2020
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Received 27 Dec, 2020
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Received 19 Dec, 2020
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Received 01 Dec, 2020
Reviewer #3 agreed
On 30 Nov, 2020
Reviewer #2 agreed
On 29 Nov, 2020
Reviewer #1 agreed
On 24 Nov, 2020
Reviewers invited
Invitations sent on 21 Nov, 2020
Editor assigned
On 18 Nov, 2020
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On 18 Nov, 2020
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On 18 Nov, 2020
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Subject Areas
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A new preprint design is coming!
We have improved readability, clarity, accessibility, and opportunities for engagement.
See the published version of this preprint at Microbial Cell Factories.
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.
Learn more about In Review
Research
Weifeng Liu
Institute of Microbiology Chinese Academy of Sciences
Corresponding Author
ORCiD: https://orcid.org/0000-0001-7218-3932
Badges
Prescreen
This manuscript passed our Prescreen assessment
Complete author information
Appropriate declaration statements
Potential risks to human health
Prescreen
Peer Review Timeline
CURRENT STATUS: Published
View the published article at Microbial Cell Factories.
Version 1
Posted 01 Dec, 2020
No community comments so far
Editorial decision: Major Revision
On 28 Dec, 2020
Review #1 received
Received 27 Dec, 2020
Review #3 received
Received 19 Dec, 2020
Review #2 received
Received 01 Dec, 2020
Reviewer #3 agreed
On 30 Nov, 2020
Reviewer #2 agreed
On 29 Nov, 2020
Reviewer #1 agreed
On 24 Nov, 2020
Reviewers invited
Invitations sent on 21 Nov, 2020
Editor assigned
On 18 Nov, 2020
Submission checks complete
On 18 Nov, 2020
Editor invited
On 18 Nov, 2020
First submitted
On 15 Nov, 2020
Metrics
Comments: 0
PDF Downloads: ...
HTML Views: ...
Subject Areas
Applied & Industrial Microbiology
License:
This work is licensed under a CC BY 4.0 License. Read Full License
View the published article at Microbial Cell Factories.
Background
Phenylpropanoid including raspberry ketone, is a kind of important natural plant product and widely used in pharmaceuticals, chemicals, cosmetics, and healthcare products. Bioproduction of phenylpropanoid in Escherichia coli and other microbial cell factories is an attractable approach considering the low phenylpropanoid contents in plants. However, it is usually difficult to produce high titer phenylpropanoid production when fermentation using glucose as carbon source. Developing novel bioprocess using alternative sources might provide a solution to this problem. In this study, typical phenylpropanoid raspberry ketone was used as the target product to develop a biosynthesis pathway for phenylpropanoid production from fatty acids, a promising alternative low-cost feedstock.
Results
A raspberry ketone biosynthesis module was developed and optimized by introducing 4-coumarate-CoA ligase (4CL), benzalacetone synthase (BAS), and raspberry ketone reductase (RZS) in Escherichia coli strains CR1-CR4. Then strain CR5 was developed by introducing raspberry ketone biosynthesis module into a fatty acids-utilization chassis FA09 to achieve production of raspberry ketone from fatty acids feedstock. However, the production of raspberry ketone was still limited by the low biomass and unable to substantiate whole-cell bioconversion process. Thus, a process by coordinately using fatty-acids and glycerol was developed. In addition, we systematically screened and optimized fatty acids-response promoters. The optimized promoter Pfrd3 was then successfully used for the efficient expression of key enzymes of raspberry ketone biosynthesis module during bioconversion from fatty acids. The final engineered strain CR8 could efficiently produce raspberry ketone repeatedly using bioconversion from fatty acids feedstock strategy, and about 291.3 mg/L raspberry ketone was produced.
Conclusion
Metabolically engineered Escherichia coli strains were successfully developed for raspberry ketone production from fatty acids using several strategies, including optimization of bioconversion process and fine-tuning key enzyme expression. This study provides an essential reference to establish the low-cost biological manufacture of phenylpropanoids compounds.
Figure 1
Figure 2
Figure 3
Figure 4
Figure 5
Figure 6
Figure 7
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