Journal
METABOLIC ENGINEERING
Volume 68, Issue -, Pages 199-209Publisher
ACADEMIC PRESS INC ELSEVIER SCIENCE
DOI: 10.1016/j.ymben.2021.10.006
Keywords
Photosynthesis; Cyanobacteria; Oxygen-tolerant hydrogenase; Sustainable hydrogen production
Categories
Funding
- European Regional Development Fund (EFRE) [100361842]
- means of taxation based on the budget adopted by the representatives of the Landtag of Saxony
- EFRE
- Deutsche Forschungsgemeinschaft (DFG, Germany) under Germany's Excellence Strategy [EXC 2008-390540038, 405325648]
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This study successfully addressed the challenge of oxygen release during photosynthesis-driven H-2 production by introducing an oxygen-tolerant hydrogenase into cyanobacteria. The hydrogenase enabled the bacteria to utilize H-2 to fix CO2 and produce H-2 under different conditions, laying a foundation for further engineering advancements.
Molecular hydrogen (H-2) is considered as an ideal energy carrier to replace fossil fuels in future. Biotechnological H-2 production driven by oxygenic photosynthesis appears highly promising, as biocatalyst and H-2 syntheses rely mainly on light, water, and CO2 and not on rare metals. This biological process requires coupling of the photosynthetic water oxidizing apparatus to a H-2-producing hydrogenase. However, this strategy is impeded by the simultaneous release of oxygen (O-2) which is a strong inhibitor of most hydrogenases. Here, we addressed this challenge, by the introduction of an O-2-tolerant hydrogenase into phototrophic bacteria, namely the cyanobacterial model strain Synechocystis sp. PCC 6803. To this end, the gene cluster encoding the soluble, O-2-tolerant, and NAD(H)-dependent hydrogenase from Ralstonia eutropha (ReSH) was functionally transferred to a Synechocystis strain featuring a knockout of the native O-2 sensitive hydrogenase. Intriguingly, photosynthetically active cells produced the O-2 tolerant ReSH, and activity was confirmed in vitro and in vivo. Further, ReSH enabled the constructed strain Syn_ReSH+ to utilize H-2 as sole electron source to fix CO2. Syn_ReSH+ also was able to produce H-2 under dark fermentative conditions as well as in presence of light, under conditions fostering intracellular NADH excess. These findings highlight a high level of interconnection between ReSH and cyano-bacterial redox metabolism. This study lays a foundation for further engineering, e.g., of electron transfer to ReSH via NADPH or ferredoxin, to finally enable photosynthesis-driven H-2 production.
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