title: Modular electron-transport chains from eukaryotic organelles function to support nitrogenase activity.
abstract: A large number of genes are necessary for the biosynthesis and activity of the enzyme nitrogenase to carry out the process of biological nitrogen fixation (BNF), which requires large amounts of ATP and reducing power. The multiplicity of the genes involved, the oxygen sensitivity of nitrogenase, plus the demand for energy and reducing power, are thought to be major obstacles to engineering BNF into cereal crops. Genes required for nitrogen fixation can be considered as three functional modules encoding electron-transport components (ETCs), proteins required for metal cluster biosynthesis, and the "core" nitrogenase apoenzyme, respectively. Among these modules, the ETC is important for the supply of reducing power. In this work, we have usedEscherichia colias a chassis to study the compatibility between molybdenum and the iron-only nitrogenases with ETC modules from target plant organelles, including chloroplasts, root plastids, and mitochondria. We have replaced an ETC module present in diazotrophic bacteria with genes encoding ferredoxin-NADPH oxidoreductases (FNRs) and their cognate ferredoxin counterparts from plant organelles. We observe that the FNR-ferredoxin module from chloroplasts and root plastids can support the activities of both types of nitrogenase. In contrast, an analogous ETC module from mitochondria could not function in electron transfer to nitrogenase. However, this incompatibility could be overcome with hybrid modules comprising mitochondrial NADPH-dependent adrenodoxin oxidoreductase and theAnabaenaferredoxins FdxH or FdxB. We pinpoint endogenous ETCs from plant organelles as power supplies to support nitrogenase for future engineering of diazotrophy in cereal crops.
authors.affiliation: State Key Laboratory of Protein and Plant Gene Research, College of Life Sciences, Peking University, Beijing 100871, China.
authors.forename: Jianguo
authors.initials: J
authors.lastname: Yang
authors.affiliation: State Key Laboratory of Protein and Plant Gene Research, College of Life Sciences, Peking University, Beijing 100871, China.
authors.forename: Xiaqing
authors.initials: X
authors.lastname: Xie
authors.affiliation: State Key Laboratory of Protein and Plant Gene Research, College of Life Sciences, Peking University, Beijing 100871, China.
authors.forename: Mingxuan
authors.initials: M
authors.lastname: Yang
authors.affiliation: Department of Molecular Microbiology, John Innes Centre, Norwich, NR4 7UH, United Kingdom ray.dixon@jic.ac.uk wangyp@pku.edu.cn.
authors.forename: Ray
authors.initials: R
authors.lastname: Dixon
authors.affiliation: State Key Laboratory of Protein and Plant Gene Research, College of Life Sciences, Peking University, Beijing 100871, China; ray.dixon@jic.ac.uk wangyp@pku.edu.cn.
authors.forename: Yi-Ping
authors.initials: YP
authors.lastname: Wang
mesh_terms: D017033:Anabaena / Q000201:enzymology / Q000235:genetics; D004579:Electron Transport; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism; D056890:Eukaryota / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism; D005287:Ferredoxin-NADP Reductase / Q000235:genetics / Q000378:metabolism; D008982:Molybdenum / Q000378:metabolism; D009591:Nitrogenase / Q000235:genetics / Q000378:metabolism*; D015388:Organelles / Q000201:enzymology* / Q000235:genetics; D010084:Oxidation-Reduction
chemical_list: D008982:Molybdenum; D005287:Ferredoxin-NADP Reductase; D009591:Nitrogenase
keywords: electron transport; nitrogen fixation; nitrogenase engineering; plant organelles