| 1. |
- Lindblad, Peter, et al.
(författare)
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CyanoFactory, a European consortium to develop technologies needed to advance cyanobacteria as chassis for production of chemicals and fuels
- 2019
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Ingår i: Algal Research. - : Elsevier. - 2211-9264. ; 41
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Forskningsöversikt (refereegranskat)abstract
- CyanoFactory, Design, construction and demonstration of solar biofuel production using novel (photo) synthetic cell factories, was an R&D project developed in response to the European Commission FP7-ENERGY-2012-1 call "Future Emerging Technologies" and the need for significant advances in both new science and technologies to convert solar energy into a fuel. CyanoFactory was an example of "purpose driven" research and development with identified scientific goals and creation of new technologies. The present overview highlights significant outcomes of the project, three years after its successful completion. The scientific progress of CyanoFactory involved: (i) development of a ToolBox for cyanobacterial synthetic biology; (ii) construction of DataWarehouse/Bioinformatics web-based capacities and functions; (iii) improvement of chassis growth, functionality and robustness; (iv) introduction of custom designed genetic constructs into cyanobacteria, (v) improvement of photosynthetic efficiency towards hydrogen production; (vi) biosafety mechanisms; (vii) analyses of the designed cyanobacterial cells to identify bottlenecks with suggestions on further improvements; (viii) metabolic modelling of engineered cells; (ix) development of an efficient laboratory scale photobioreactor unit; and (x) the assembly and experimental performance assessment of a larger (1350 L) outdoor flat panel photobioreactor system during two seasons. CyanoFactory - Custom design and purpose construction of microbial cells for the production of desired products using synthetic biology - aimed to go beyond conventional paths to pursue innovative and high impact goals. CyanoFactory brought together ten leading European partners (universities, research organizations and enterprises) with a common goal - to develop the future technologies in Synthetic biology and Advanced photobioreactors.
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| 2. |
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| 3. |
- Wegelius, Adam, et al.
(författare)
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Design and characterization of a synthetic minimal promoter for heterocyst-specific expression in filamentous cyanobacteria
- 2018
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Ingår i: PLOS ONE. - : Public Library of Science (PLoS). - 1932-6203. ; 13:9
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Tidskriftsartikel (refereegranskat)abstract
- Short and well defined promoters are essential for advancing cyanobacterial biotechnology. The heterocyst of Nostoc sp. is suggested as a microbial cell factory for oxygen sensitive catalysts, such as hydrogenases for hydrogen production, due to its microoxic environment. We identified and predicted promoter elements of possible significance through a consensus strategy using a pool of heterocyst-induced DIF+ promoters known from Anabaena sp. PCC 7120. To test if these conserved promoter elements were crucial for heterocyst-specific expression, promoter-yfp reporter constructs were designed. The characterization was accomplished by replacing, -35 and -10 regions and the upstream element, with well described elements from the trc promoter of Escherichia coli, which is also functional in Nostoc sp. From the in vivo spatial fluorescence of the different promoter-yfp reporters in Nostoc punctiforme ATCC 29133, we concluded that both the consensus -35 and extended -10 regions were important for heterocyst-specific expression. Further that the promoter strength could be improved by the addition of an upstream element. We designed a short synthetic promoter of 48 nucleotides, PsynDIF, including a consensus DIF1 sequence, a 17 base pair stretch of random nucleotides and an extended consensus -10 region, and thus generated the shortest promoter for heterocyst-specific expression to date.
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| 4. |
- Wegelius, Adam, 1989-
(författare)
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Expression and synthetic activation of [FeFe]-hydrogenases in cyanobacteria
- 2020
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Photosynthetic microbes can be utilized for hydrogen production, generating a clean, carbon neutral energy carrier from abundant substrates. Cyanobacteria are photosynthetic prokaryotes with large potential for biotechnological energy applications and several strains are capable of hydrogen production. This production is catalysed by a bi-directional [NiFe]-hydrogenase, or by nitrogenase during nitrogen fixation. However, nature’s foremost hydrogen producing enzymes, the [FeFe]-hydrogenases, are not present in these organisms. Many [FeFe]-hydrogenases boast incredible catalytic activities and high bias towards proton reduction. Introduction of a suitable [FeFe]-hydrogenase in a cyanobacterial host could greatly improve the hydrogen production capacity. Unfortunately, generation and characterisation of cyanobacterial strains carrying active [FeFe]-hydrogenases is stalled by the intricate maturation process associated with these enzymes.In this thesis, I investigate heterologous expression and artificial maturation of [FeFe]-hydrogenases in cyanobacteria. Genetic tools to reliably express [FeFe]-hydrogenases were developed and tested in the unicellular cyanobacterium Synechocystis PCC 6083, and in heterocysts of the filamentous cyanobacterium Nostoc punctiforme ATCC 29133. Following heterologous expression, functional, semisynthetic [FeFe]-hydrognases operating in vivo in cyanobacterial cells were generated by synthetic activation. The procedure proved successful in both the unicellular and filamentous strain, and for [FeFe]-hydrogenases from different groups and subclasses. The semisynthetic enzymes proved capable of hydrogen production under different environmental conditions and links to the metabolism of the host cell. Hydrogen production capacity proved long-lived and was retained for several days. In Nostoc punctiforme, synthetic activation was confirmed to generate active [FeFe]-hydrogenase in both vegetative cells and heterocyst.The results presented in this thesis demonstrate a novel way to explore in vivo hydrogen production from heterologous [FeFe]-hydrogenases in cyanobacteria. In the search for suitable candidates for H2 production systems, synthetic activation may be used to investigate a wide range of [FeFe]-hydrogenases, strains and cultivation conditions, circumventing the need of elaborate maturation machinery optimisation.
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| 5. |
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| 6. |
- Wegelius, Adam, et al.
(författare)
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Generation of a functional, semisynthetic [FeFe]-hydrogenase in a photosynthetic microorganism
- 2018
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Ingår i: Energy & Environmental Science. - : Royal Society of Chemistry (RSC). - 1754-5692 .- 1754-5706. ; 11:11, s. 3163-3167
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Tidskriftsartikel (refereegranskat)abstract
- [FeFe]-Hydrogenases are hydrogen producing metalloenzymes with excellent catalytic capacities, highly relevant in the context of a future hydrogen economy. Here we demonstrate the synthetic activation of a heterologously expressed [FeFe]-hydrogenase in living cells of Synechocystis PCC 6803, a photoautotrophic microbial chassis with high potential for biotechnological energy applications. H-2-Evolution assays clearly show that the non-native, semi-synthetic enzyme links to the native metabolism in living cells.
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| 7. |
- Wegelius, Adam, et al.
(författare)
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Semisynthetic [FeFe]-hydrogenase with stable expression and H2 production capacity in a photosynthetic microbe
- 2021
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Ingår i: Cell Reports Physical Science. - : Elsevier. - 2666-3864. ; 2:3
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Tidskriftsartikel (refereegranskat)abstract
- Hydrogen (H2) is a promising future chemical energy carrier and feedstock with several renewable production options, including electrolyzers and biological/bioinspired systems. The top H2 producers in nature are [FeFe]-hydrogenases, high turnover metalloenzymes with a complex maturation process that can be circumvented by artificial synthetic activation. Here, we report the expression and activation of group A and D [FeFe]-hydrogenases in a photosynthetic host organism, the unicellular cyanobacterium Synechocystis PCC 6803. The hydrogenase from Solobacterium moorei (group A) facilitates high in vivo H2 production from purely photoautotrophically generated substrates and unmistakably links to the metabolism of the photosynthetic host. Cells harboring the non-native, semisynthetic enzyme retain their H2 production capacity for several days after synthetic activation. This work expands both the number and the diversity of [FeFe]-hydrogenases examined in a photosynthetic background and provides important insights for future investigations into the development and understanding of biological and biohybrid H2 production systems.
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