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Background: Oleaginous microalgae represent a valuable resource for the production of high value molecules. Considering the importance of omega-3 long chain polyunsaturated fatty acids (LC-PUFAs) for human health and nutrition the yields of high value eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) require significant improvement to meet demand, however, the current cost of production remains high. A promising approach is to metabolically engineer strains with enhanced levels of triacylglycerols (TAGs) enriched in EPA and DHA.
Results: Recently we have engineered the marine diatom Phaeodactylum tricornutum to accumulate enhanced levels of DHA in TAG. To further improve the incorporation of omega-3 LC-PUFAs in TAG we focused our effort on the identification of a type 2 acyl-CoA:diacylglycerol acyltransferase (DGAT) capable of improving lipid production and the incorporation of DHA in TAG. DGAT is a key enzyme in lipid synthesis. Following a diatom based in vivo screen of candidate DGATs, a native P. tricornutum DGAT2B was taken forward for detailed characterisation. Overexpression of the endogenous P. tricornutum DGAT2B was confirmed by qRT-PCR and the transgenic strain grew successfully in comparison to wildtype. PtDGAT2B has broad substrate specificity with preferences for C16 and LC-PUFAs acyl groups. Moreover, the overexpression of an endogenous DGAT2B resulted in higher lipid yields and enhanced levels of DHA in TAG. Furthermore, a combined overexpression of the endogenous DGAT2B and ectopic expression of a Δ5-elongase showed how iterative metabolic engineering can be used to increase DHA and TAG content, irrespective of nitrogen treatment.
Conclusion: This study provides further insight into lipid metabolism in P. tricornutum and suggests a metabolic engineering approach for the efficient production of EPA and DHA in microalgae.
Keywords
Acyl-CoA:diacylglycerol acyltransferase (DGAT), eicosapentaenoic acid, docosahexaenoic acid, triacylgycerol, omega-3 fatty acids, Phaeodactylum tricornutum.
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View the published article at Biotechnology for Biofuels.
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On 07 May, 2020
Editorial decision: Accept
On 07 May, 2020
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On 06 May, 2020
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On 06 May, 2020
No comments provided
Editorial decision: Minor revision
On 09 Apr, 2020
Review #2 received
Received 08 Apr, 2020
Review #4 received
Received 27 Mar, 2020
Review #3 received
Received 27 Mar, 2020
Reviewer #4 agreed
On 26 Mar, 2020
Reviewer #3 agreed
On 20 Mar, 2020
Review #1 received
Received 29 Jan, 2020
Reviewer #2 agreed
On 22 Jan, 2020
Reviewer #1 agreed
On 20 Jan, 2020
Reviewers invited
Invitations sent on 19 Jan, 2020
First submitted
On 17 Jan, 2020
Editor assigned
On 17 Jan, 2020
Submission checks complete
On 16 Jan, 2020
Editor invited
On 16 Jan, 2020
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See the published version of this preprint at Biotechnology for Biofuels.
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
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 Biotechnology for Biofuels.
Version 2
Posted 19 May, 2020
No community comments so far
Editor assigned
On 07 May, 2020
Editorial decision: Accept
On 07 May, 2020
Submission checks complete
On 06 May, 2020
Editor invited
On 06 May, 2020
No comments provided
Editorial decision: Minor revision
On 09 Apr, 2020
Review #2 received
Received 08 Apr, 2020
Review #4 received
Received 27 Mar, 2020
Review #3 received
Received 27 Mar, 2020
Reviewer #4 agreed
On 26 Mar, 2020
Reviewer #3 agreed
On 20 Mar, 2020
Review #1 received
Received 29 Jan, 2020
Reviewer #2 agreed
On 22 Jan, 2020
Reviewer #1 agreed
On 20 Jan, 2020
Reviewers invited
Invitations sent on 19 Jan, 2020
First submitted
On 17 Jan, 2020
Editor assigned
On 17 Jan, 2020
Submission checks complete
On 16 Jan, 2020
Editor invited
On 16 Jan, 2020
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
View the published article at Biotechnology for Biofuels.
Background: Oleaginous microalgae represent a valuable resource for the production of high value molecules. Considering the importance of omega-3 long chain polyunsaturated fatty acids (LC-PUFAs) for human health and nutrition the yields of high value eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) require significant improvement to meet demand, however, the current cost of production remains high. A promising approach is to metabolically engineer strains with enhanced levels of triacylglycerols (TAGs) enriched in EPA and DHA.
Results: Recently we have engineered the marine diatom Phaeodactylum tricornutum to accumulate enhanced levels of DHA in TAG. To further improve the incorporation of omega-3 LC-PUFAs in TAG we focused our effort on the identification of a type 2 acyl-CoA:diacylglycerol acyltransferase (DGAT) capable of improving lipid production and the incorporation of DHA in TAG. DGAT is a key enzyme in lipid synthesis. Following a diatom based in vivo screen of candidate DGATs, a native P. tricornutum DGAT2B was taken forward for detailed characterisation. Overexpression of the endogenous P. tricornutum DGAT2B was confirmed by qRT-PCR and the transgenic strain grew successfully in comparison to wildtype. PtDGAT2B has broad substrate specificity with preferences for C16 and LC-PUFAs acyl groups. Moreover, the overexpression of an endogenous DGAT2B resulted in higher lipid yields and enhanced levels of DHA in TAG. Furthermore, a combined overexpression of the endogenous DGAT2B and ectopic expression of a Δ5-elongase showed how iterative metabolic engineering can be used to increase DHA and TAG content, irrespective of nitrogen treatment.
Conclusion: This study provides further insight into lipid metabolism in P. tricornutum and suggests a metabolic engineering approach for the efficient production of EPA and DHA in microalgae.
Figure 1
Figure 2
Figure 3
Figure 4
Figure 5
Figure 6
Figure 7
Figure 8
This is a list of supplementary files associated with this preprint. Click to download.
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