Plant Biochemistry, Faculty of Biology & Biotechnology, Ruhr University Bochum, 44780 Bochum, Germany.
Photobiotechnology, Faculty of Biology & Biotechnology, Ruhr University Bochum, 44780 Bochum, Germany.
Biochim Biophys Acta Bioenerg. 2018 Apr;1859(4):253-262. doi: 10.1016/j.bbabio.2018.01.006. Epub 2018 Jan 31.
Utilization of electrons from the photosynthetic water splitting reaction for the generation of biofuels, commodities as well as application in biotransformations requires a partial rerouting of the photosynthetic electron transport chain. Due to its rather negative redox potential and its bifurcational function, ferredoxin at the acceptor side of Photosystem 1 is one of the focal points for such an engineering. With hydrogen production as model system, we show here the impact and potential of redox partner design involving ferredoxin (Fd), ferredoxin-oxido-reductase (FNR) and [FeFe]‑hydrogenase HydA1 on electron transport in a future cyanobacterial design cell of Synechocystis PCC 6803. X-ray-structure-based rational design and the allocation of specific interaction residues by NMR-analysis led to the construction of Fd- and FNR-mutants, which in appropriate combination enabled an about 18-fold enhanced electron flow from Fd to HydA1 (in competition with equimolar amounts of FNR) in in vitro assays. The negative impact of these mutations on the Fd-FNR electron transport which indirectly facilitates H production (with a contribution of ≤42% by FNR variants and ≤23% by Fd-variants) and the direct positive impact on the Fd-HydA1 electron transport (≤23% by Fd-mutants) provide an excellent basis for the construction of a hydrogen-producing design cell and the study of photosynthetic efficiency-optimization with cyanobacteria.
利用光合作用水分解反应中的电子来生成生物燃料、商品，并将其应用于生物转化，需要对光合作用电子传递链进行部分改道。由于铁氧还蛋白具有相当负的氧化还原电位和分叉功能，因此它是光合作用 1 号系统接受体侧的焦点之一，可用于此类工程设计。以产氢作为模型系统，我们在此展示了涉及铁氧还蛋白（Fd）、铁氧还蛋白氧化还原酶（FNR）和 [FeFe]-氢化酶 HydA1 的氧化还原伴侣设计对未来设计的集胞藻 PCC 6803 中蓝藻设计细胞电子传递的影响和潜力。基于 X 射线结构的合理设计和通过 NMR 分析分配特定相互作用残基，构建了 Fd 和 FNR 突变体，在适当的组合下，在体外测定中，它们使电子从 Fd 流向 HydA1 的流量增加了约 18 倍（与 FNR 的等摩尔量竞争）。这些突变对 Fd-FNR 电子传递的负面影响（间接促进 H 生成，其中 FNR 变体的贡献≤42%，Fd 变体的贡献≤23%）和对 Fd-HydA1 电子传递的直接积极影响（Fd 突变体的贡献≤23%）为构建产氢设计细胞和研究蓝藻的光合效率优化提供了极好的基础。
