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Filipe Branco dos Santos
University of Amsterdam Swammerdam Institute for Life Sciences
Corresponding Author
ORCiD: https://orcid.org/0000-0002-4268-8080
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Background Mannitol is a C(6) polyol that is used in the food and medical sector as a sweetener and antioxidant, respectively. The sustainable production of mannitol, especially via the direct conversion of CO 2 by photosynthetic cyanobacteria, has become increasingly appealing. However, previous work aiming to achieve mannitol production in the marine Synechococcus sp. PCC 7002 via heterologous expression of mannitol-1-phosphate-5-dehydrogenase ( mtlD ) and mannitol-1-phosphatase ( m1p , in short: a ‘mannitol cassette’), proved to be genetically unstable.
Results Here, we explore the stabilizing effect that mannitol production may have on cells faced with osmotic stress, in the freshwater cyanobacterium Synechocystis sp. PCC 6803. We first validated that mannitol can function as a compatible solute in Synechocystis sp. PCC 6803, and in derivative strains in which the ability to produce one or both of the native compatible solutes was impaired. Wild type Synechocystis , complemented with a mannitol cassette, indeed showed increased salt tolerance, which was even more evident in Synechocystis strains in which the ability to synthesize the endogenous compatible solutes was impaired. Next we tested the genetic stability of all these strains with respect to their mannitol productivity, with and without salt stress, during prolonged turbidostat cultivations. The obtained results show that mannitol production under salt stress conditions in the Synechocystis strain that cannot synthesize its endogenous compatible solutes is remarkably stable, while the control strain completely loses this ability in only 6 days. DNA sequencing results of the control groups that lost the ability to synthesize mannitol revealed that multiple types of mutation occurred in the mtlD gene that can explain the disruption of mannitol production.
Conclusions Mannitol production in freshwater Synechocsytis sp. PCC6803 confers it with increased salt tolerance. Under this strategy, genetically instability which was the major challenge for mannitol production in cyanobacteria is tackled. This paper marks the first report of utilization of the response to salt stress as a factor that can increase the stability of mannitol production in a cyanobacterial cell factory.
Keywords
(D-)Mannitol, Synechocystis sp. PCC 6803, Compatible solutes, Production stability,Salt stress
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CURRENT STATUS: Published
View the published article at Biotechnology for Biofuels and Bioproducts.
No community comments so far
Editorial decision: Accept
On 22 Jun, 2020
Editor assigned
On 20 Jun, 2020
Submission checks complete
On 19 Jun, 2020
Editor invited
On 19 Jun, 2020
Version 2
Posted 31 May, 2020
No comments provided
Editorial decision: Minor revision
On 07 Jun, 2020
Review #3 received
Received 01 Jun, 2020
Review #1 received
Received 27 May, 2020
Review #2 received
Received 27 May, 2020
Reviewer #3 agreed
On 26 May, 2020
Reviewer #2 agreed
On 24 May, 2020
Reviewer #1 agreed
On 23 May, 2020
Reviewers invited
Invitations sent on 22 May, 2020
Editor assigned
On 22 May, 2020
Submission checks complete
On 21 May, 2020
Editor invited
On 21 May, 2020
No comments provided
Review #1 received
Received 20 Apr, 2020
Editorial decision: Major revision
On 20 Apr, 2020
Review #3 received
Received 03 Apr, 2020
Review #2 received
Received 03 Apr, 2020
Reviewer #3 agreed
On 26 Mar, 2020
Reviewer #2 agreed
On 20 Mar, 2020
Reviewer #1 agreed
On 17 Mar, 2020
Reviewers invited
Invitations sent on 16 Mar, 2020
Editor invited
On 14 Mar, 2020
Editor assigned
On 14 Mar, 2020
First submitted
On 13 Mar, 2020
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Subject Areas
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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
Filipe Branco dos Santos
University of Amsterdam Swammerdam Institute for Life Sciences
Corresponding Author
ORCiD: https://orcid.org/0000-0002-4268-8080
Badges
Prescreen
This manuscript passed our Prescreen assessment
Complete author information
Appropriate declaration statements
Potential risks to human health
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Peer Review Timeline
CURRENT STATUS: Published
View the published article at Biotechnology for Biofuels and Bioproducts.
No community comments so far
Editorial decision: Accept
On 22 Jun, 2020
Editor assigned
On 20 Jun, 2020
Submission checks complete
On 19 Jun, 2020
Editor invited
On 19 Jun, 2020
Version 2
Posted 31 May, 2020
No comments provided
Editorial decision: Minor revision
On 07 Jun, 2020
Review #3 received
Received 01 Jun, 2020
Review #1 received
Received 27 May, 2020
Review #2 received
Received 27 May, 2020
Reviewer #3 agreed
On 26 May, 2020
Reviewer #2 agreed
On 24 May, 2020
Reviewer #1 agreed
On 23 May, 2020
Reviewers invited
Invitations sent on 22 May, 2020
Editor assigned
On 22 May, 2020
Submission checks complete
On 21 May, 2020
Editor invited
On 21 May, 2020
No comments provided
Review #1 received
Received 20 Apr, 2020
Editorial decision: Major revision
On 20 Apr, 2020
Review #3 received
Received 03 Apr, 2020
Review #2 received
Received 03 Apr, 2020
Reviewer #3 agreed
On 26 Mar, 2020
Reviewer #2 agreed
On 20 Mar, 2020
Reviewer #1 agreed
On 17 Mar, 2020
Reviewers invited
Invitations sent on 16 Mar, 2020
Editor invited
On 14 Mar, 2020
Editor assigned
On 14 Mar, 2020
First submitted
On 13 Mar, 2020
Submission checks complete
On 13 Mar, 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 and Bioproducts.
Background Mannitol is a C(6) polyol that is used in the food and medical sector as a sweetener and antioxidant, respectively. The sustainable production of mannitol, especially via the direct conversion of CO 2 by photosynthetic cyanobacteria, has become increasingly appealing. However, previous work aiming to achieve mannitol production in the marine Synechococcus sp. PCC 7002 via heterologous expression of mannitol-1-phosphate-5-dehydrogenase ( mtlD ) and mannitol-1-phosphatase ( m1p , in short: a ‘mannitol cassette’), proved to be genetically unstable.
Results Here, we explore the stabilizing effect that mannitol production may have on cells faced with osmotic stress, in the freshwater cyanobacterium Synechocystis sp. PCC 6803. We first validated that mannitol can function as a compatible solute in Synechocystis sp. PCC 6803, and in derivative strains in which the ability to produce one or both of the native compatible solutes was impaired. Wild type Synechocystis , complemented with a mannitol cassette, indeed showed increased salt tolerance, which was even more evident in Synechocystis strains in which the ability to synthesize the endogenous compatible solutes was impaired. Next we tested the genetic stability of all these strains with respect to their mannitol productivity, with and without salt stress, during prolonged turbidostat cultivations. The obtained results show that mannitol production under salt stress conditions in the Synechocystis strain that cannot synthesize its endogenous compatible solutes is remarkably stable, while the control strain completely loses this ability in only 6 days. DNA sequencing results of the control groups that lost the ability to synthesize mannitol revealed that multiple types of mutation occurred in the mtlD gene that can explain the disruption of mannitol production.
Conclusions Mannitol production in freshwater Synechocsytis sp. PCC6803 confers it with increased salt tolerance. Under this strategy, genetically instability which was the major challenge for mannitol production in cyanobacteria is tackled. This paper marks the first report of utilization of the response to salt stress as a factor that can increase the stability of mannitol production in a cyanobacterial cell factory.
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.
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