| 1. |
- Agervald, Åsa, et al.
(författare)
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CalA, a cyanobacterial AbrB protein, interacts with the upstream region of hypC and acts as a repressor of its transcription in the cyanobacterium Nostoc sp. strain PCC 7120
- 2010
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Ingår i: Applied and Environmental Microbiology. - 0099-2240 .- 1098-5336. ; 76:3, s. 880-890
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Tidskriftsartikel (refereegranskat)abstract
- The filamentous, heterocystous, nitrogen-fixing cyanobacterium Nostoc sp. strain PCC 7120 may contain, depending on growth condition, up to two hydrogenases directly involved in hydrogen metabolism. HypC is one out of at least seven auxiliary gene products required for synthesis of a functional hydrogenase, specifically involved in the maturation of the large subunit. In this study we present a protein, Alr0946, belonging to the transcription regulator family AbrB, which in protein-DNA assays was found to interact with the upstream region of hypC. Transcriptional investigations showed that alr0946 is co-transcribed with the downstream gene alr0947, which encodes a putative protease from the abortive infection superfamily, Abi. Alr0946 was shown to interact specifically not only with the upstream region of hypC but also with its own upstream region, acting as a repressor on both. The bidirectional hydrogenase activity was significant down-regulated when Alr0946 was over-expressed demonstrating a correlation to the transcription factor, either direct or indirect. In silico studies showed that homologues to both Alr0946 and Alr0947 are highly conserved proteins within cyanobacteria with a very similar physical organisation of the corresponding structural genes. Possible functions of the co-transcribed downstream protein Alr0947 are presented. In addition, we present a 3D model of the CyAbrB domain of Alr0946 and putative DNA-binding mechanisms are discussed.
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| 2. |
- Agervald, Åsa, et al.
(författare)
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CRISPR in the extended hyp-operon of the cyanobacterium Nostoc sp. strain PCC 7120, characteristics and putative function(s)
- 2012
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Ingår i: International journal of hydrogen energy. - : Elsevier BV. - 0360-3199 .- 1879-3487. ; 37:10, s. 8828-8833
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Tidskriftsartikel (refereegranskat)abstract
- The presence of small RNAs (sRNA) and their functions in transcriptional regulation has lately turned into a hot topic. Since cyanobacteria often face changes in the surrounding environment, they need to have a well working system for stress response. Quick adaption is necessary, and an RNA-based regulatory system is thus useful. One example of these sRNAs is CRISPRs. In this work we report the existence of a CRISPR within the hyp-operon (hyp genes encode proteins responsible for the maturation of hydrogenases) of the filamentous cyanobacterium Nostoc sp. strain PCC 7120. We present data concerning its characteristics and putative function(s) and raise the question concerning the importance of this CRISPR array and other CRISPR systems in general. In addition, we discuss the use of the CRISPR system as a potential bacterial genetic defence mechanism to achieve robust, cyanobacterial cultures in large scale, commercial production units.
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| 3. |
- Agervald, Åsa
(författare)
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Maturation and Regulation of Cyanobacterial Hydrogenases
- 2009
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Accelerated global warming plus an increasing need for energy is an equation not easily solved, thus new forms of sustainable energy production are urgently requested. In this context hydrogen production based on a cyanobacterial system offers an environmentally friendly alternative for energy capture and conversion. Cyanobacteria can produce hydrogen gas from sun light and water through the combination of photosystems and hydrogenases, and are suitable to cultivate in large scale. In the present thesis the maturation process of [NiFe]-hydrogenases is investigated with special focus on transcription of the accessory genes encoding proteins needed for assembly of the large and possibly also for the small hydrogenase subunit. The cyanobacteria used are two N2-fixing, filamentous, heterocystous strains; Nostoc sp. strain PCC 7120 and Nostoc punctiforme PCC 73102. For a biotechnological exploration of hydrogen production tools for regulatory purposes are important. The transcription factor CalA (cyanobacterial AbrB like) (Alr0946 in the genome) in Nostoc sp. strain PCC 7120 was found to be involved in hydrogen metabolism by regulating the transcription of the maturation protein HypC. Further the bidirectional hydrogenase activity was down-regulated in the presence of elevated levels of CalA, a result important to take into account when optimizing cyanobacteria for hydrogen production. CalA regulates at least 25 proteins in Nostoc sp. strain PCC 7120 and one of the down-regulated proteins was superoxide dismutase, FeSOD. The characterization of FeSOD shows that it has a specific and important function in the oxidative stress tolerance of Nostoc sp. stain PCC 7120. Since CalA is involved in regulation of both the hydrogen metabolism as well as stress responses these findings indicate that Alr0946 is an important transcription factor in Nostoc sp. strain PCC 7120 active on a global level in the cell. This thesis adds more knowledge concerning maturation and regulation of cyanobacterial hydrogenases which might be useful for future large scale hydrogen.
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| 4. |
- Agervald, Åsa, et al.
(författare)
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The CyAbrB transcription factor Alr0946 regulates the iron superoxide dismutase in Nostoc sp. strain PCC 7120
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Annan publikation (övrigt vetenskapligt/konstnärligt)abstract
- In the present investigation we analyse the results of induced over-production of the CyAbrB transcription factor Alr0946 in the cyanobacterium Nostoc sp. PCC 7120 with special focus on its effects on FeSOD. With gel based quantitative proteomics the induced over-expression of Alr0946 was shown to influence the abundance of at least 25 proteins. One of the proteins with a significant lower abundance was FeSOD, one of two types of superoxide dismutases in Nostoc sp. PCC 7120. The change in protein abundance was also followed by lower transcript as well as activity levels. Purified Alr0946 from Nostoc sp. PCC 7120 was shown to interact with the promoter region of alr2938, encoding FeSOD, indicating a transcriptional regulation of FeSOD by Alr0946. The Alr0946 over-expression strain showed a bleaching phenotype with lower growth rate and truncated filaments already two days after induction of over-expression. The phenotype was even more pronounced when illumination was increased from 35 to 125 μmol m-2s-1. This is in line with an increased need of FeSOD during a stronger oxidative stress. The results indicate that Alr0946 is involved in regulation of stress responses and that FeSOD has a specific and important function in the oxidative stress tolerance of the multicellular cyanobacterium Nostoc sp. PCC 7120.
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| 5. |
- Agervald, Åsa, et al.
(författare)
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The CyAbrB transcription factor CalA regulates the iron superoxide dismutase in Nostoc sp. strain PCC 7120
- 2010
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Ingår i: Environmental Microbiology. - : Wiley. - 1462-2912 .- 1462-2920. ; 12:10, s. 2826-2837
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Tidskriftsartikel (refereegranskat)abstract
- P>In the present investigation the results of induced over-production of the CyAbrB transcription factor CalA (Cyanobacterial AbrB-like, annotated as Alr0946) in the cyanobacterium Nostoc sp. PCC 7120 were analysed. The CalA overexpression strain showed a bleaching phenotype with lower growth rate and truncated filaments 2 days after induction of overexpression. The phenotype was even more pronounced when illumination was increased from 35 to 125 mu mol m-2 s-1. Using gel-based quantitative proteomics, the induced overexpression of CalA was shown to downregulate the abundance of FeSOD, one of two types of superoxide dismutases in Nostoc sp. PCC 7120. The change in protein abundance was also accompanied by lower transcript as well as activity levels. Purified recombinant CalA from Nostoc sp. PCC 7120 was shown to interact with the promoter region of alr2938, encoding FeSOD, indicating a transcriptional regulation of FeSOD by CalA. The bleaching phenotype is in line with a decreased tolerance against oxidative stress and indicates that CalA is involved in regulation of cellular responses in which FeSOD has an important and specific function in the filamentous cyanobacterium Nostoc sp. PCC 7120.
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| 6. |
- Agervald, Åsa, et al.
(författare)
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Transcription of the extended hyp-operon in Nostoc sp. strain PCC 7120
- 2008
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Ingår i: BMC Microbiology. - : Springer Science and Business Media LLC. - 1471-2180. ; 8, s. 69-
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Tidskriftsartikel (refereegranskat)abstract
- Background: The maturation of hydrogenases into active enzymes is a complex process and e. g. a correctly assembled active site requires the involvement of at least seven proteins, encoded by hypABCDEF and a hydrogenase specific protease, encoded either by hupW or hoxW. The N2fixing cyanobacterium Nostoc sp. strain PCC 7120 may contain both an uptake and a bidirectional hydrogenase. The present study addresses the presence and expression of hypgenes in Nostoc sp. strain PCC 7120. Results: RTPCRs demonstrated that the six hypgenes together with one ORF may be transcribed as a single operon. Transcriptional start points (TSPs) were identified 280 bp upstream from hypF and 445 bp upstream of hypC, respectively, demonstrating the existence of several transcripts. In addition, five upstream ORFs located in between hupSL, encoding the small and large subunits of the uptake hydrogenase, and the hypoperon, and two downstream ORFs from the hypgenes were shown to be part of the same transcript unit. A third TSP was identified 45 bp upstream of asr0689, the first of five ORFs in this operon. The ORFs are annotated as encoding unknown proteins, with the exception of alr0692 which is identified as a NifUlike protein. Orthologues of the four ORFs asr0689alr0692, with a highly conserved genomic arrangement positioned between hupSL, and the hyp genes are found in several other N2fixing cyanobacteria, but are absent in non N2fixing cyanobacteria with only the bidirectional hydrogenase. Short conserved sequences were found in six intergenic regions of the extended hypoperon, appearing between 11 and 79 times in the genome. Conclusion: This study demonstrated that five ORFs upstream of the hypgene cluster are cotranscribed with the hypgenes, and identified three TSPs in the extended hypgene cluster in Nostoc sp. strain PCC 7120. This may indicate a function related to the assembly of a functional uptake hydrogenase, hypothetically in the assembly of the small subunit of the enzyme.
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| 7. |
- Bourgade, Barbara, et al.
(författare)
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Synthetic biology in marine cyanobacteria : Advances and challenges
- 2022
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Ingår i: Frontiers in Microbiology. - : Frontiers Media S.A.. - 1664-302X. ; 13
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Forskningsöversikt (refereegranskat)abstract
- The current economic and environmental context requests an accelerating development of sustainable alternatives for the production of various target compounds. Biological processes offer viable solutions and have gained renewed interest in the recent years. For example, photosynthetic chassis organisms are particularly promising for bioprocesses, as they do not require biomass-derived carbon sources and contribute to atmospheric CO2 fixation, therefore supporting climate change mitigation. Marine cyanobacteria are of particular interest for biotechnology applications, thanks to their rich diversity, their robustness to environmental changes, and their metabolic capabilities with potential for therapeutics and chemicals production without requiring freshwater. The additional cyanobacterial properties, such as efficient photosynthesis, are also highly beneficial for biotechnological processes. Due to their capabilities, research efforts have developed several genetic tools for direct metabolic engineering applications. While progress toward a robust genetic toolkit is continuously achieved, further work is still needed to routinely modify these species and unlock their full potential for industrial applications. In contrast to the understudied marine cyanobacteria, genetic engineering and synthetic biology in freshwater cyanobacteria are currently more advanced with a variety of tools already optimized. This mini-review will explore the opportunities provided by marine cyanobacteria for a greener future. A short discussion will cover the advances and challenges regarding genetic engineering and synthetic biology in marine cyanobacteria, followed by a parallel with freshwater cyanobacteria and their current genetic availability to guide the prospect for marine species.
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| 8. |
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| 9. |
- Camsund, Daniel, et al.
(författare)
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A HupS-GFP fusion protein demonstrates a heterocyst-specific localization of the uptake hydrogenase in Nostoc punctiforme
- 2011
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Ingår i: FEMS Microbiology Letters. - : John Wiley & Sons. - 0378-1097 .- 1574-6968. ; 316:2, s. 152-159
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Tidskriftsartikel (refereegranskat)abstract
- All diazotrophic filamentous cyanobacteria contain an uptake hydrogenase that is involved in the reoxidation of H-2 produced during N-2-fixation. In Nostoc punctiforme ATCC 29133, N-2-fixation takes place in the microaerobic heterocysts, catalysed by a nitrogenase. Although the function of the uptake hydrogenase may be closely connected to that of nitrogenase, the localization in cyanobacteria has been under debate. Moreover, the subcellular localization is not understood. To investigate the cellular and subcellular localization of the uptake hydrogenase in N. punctiforme, a reporter construct consisting of the green fluorescent protein (GFP) translationally fused to HupS, within the complete hupSL operon, was constructed and transferred into N. punctiforme on a self-replicative vector by electroporation. Expression of the complete HupS-GFP fusion protein was confirmed by Western blotting using GFP antibodies. The N. punctiforme culture expressing HupS-GFP was examined using laser scanning confocal microscopy, and fluorescence was exclusively detected in the heterocysts. Furthermore, the fluorescence in mature heterocysts was localized to several small or fewer large clusters, which indicates a specificity of the subcellular localization of the uptake hydrogenase.
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| 10. |
- Cardona, Tanai, 1983-, et al.
(författare)
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Electron transfer protein complexes in the thylakoid membranes of heterocysts from the cyanobacterium Nostoc punctiforme
- 2009
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Ingår i: Biochimica et Biophysica Acta - Bioenergetics. - : Elsevier. - 0005-2728 .- 1879-2650. ; 1787:4, s. 252-263
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Tidskriftsartikel (refereegranskat)abstract
- Filamentous, heterocystous cyanobacteria are capable of nitrogen fixation and photoautotrophic growth. Nitrogen fixation takes place in heterocysts that differentiate as a result of nitrogen starvation. Heterocysts uphold a microoxic environment to avoid inactivation of nitrogenase, e.g. by downregulation of oxygenic photosynthesis. The ATP and reductant requirement for the nitrogenase reaction is considered to depend on Photosystem I, but little is known about the organization of energy converting membrane proteins in heterocysts. We have investigated the membrane proteome of heterocysts from nitrogen fixing filaments of Nostoc punctiforme sp. PCC 73102, by 2D gel electrophoresis and mass spectrometry. The membrane proteome was found to be dominated by the Photosystem I and ATP-synthase complexes.We could identify asignificant amount of assembled Photosystem II complexes containing the D1, D2, CP43, CP47 and PsbO proteins from these complexes. We could also measure light-driven in vitro electron transfer from Photosystem II in heterocyst thylakoid membranes. We did not find any partially disassembled PhotosystemII complexes lacking the CP43 protein. Several subunits of the NDH-1 complex were also identified. The relative amount of NDH-1M complexes was found to be higher than NDH-1L complexes, which might suggest a role for this complex in cyclic electron transfer in the heterocysts of Nostoc punctiforme.
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