title: A single amino acid residue controls ROS production in the respiratory Complex I from Escherichia coli.
abstract: Reactive oxygen species (ROS) production by respiratory Complex I from Escherichia coli was studied in bacterial membrane fragments and in the isolated and purified enzyme, either solubilized or incorporated in proteoliposomes. We found that the replacement of a single amino acid residue in close proximity to the nicotinamide adenine dinucleotide (NADH)-binding catalytic site (E95 in the NuoF subunit) dramatically increases the reactivity of Complex I towards dioxygen (O2 ). In the E95Q variant short-chain ubiquinones exhibit strong artificial one-electron reduction at the catalytic site, also leading to a stronger increase in ROS production. Two mechanisms can contribute to the observed kinetic effects: (a) a change in the reactivity of flavin mononucleotide (FMN) towards dioxygen at the catalytic site, and (b) a change in the population of the ROS-generating state. We propose the existence of two (closed and open) states of the NAD(+) -bound enzyme as one feature of the substrate-binding site of Complex I. The analysis of the kinetic model of ROS production allowed us to propose that the population of Complex I with reduced FMN is always low in the wild-type enzyme even at low ambient redox potentials, minimizing the rate of reaction with O2 in contrast to E95Q variant.
authors: Helsinki Bioenergetics Group, Institute of Biotechnology, University of Helsinki, PO Box 65 (Viikinkaari 1), FIN-00014, Helsinki, Finland.
authors: Juho
authors: J
authors: Knuuti
authors: Galina
authors: G
authors: Belevich
authors: Vivek
authors: V
authors: Sharma
authors: Dmitry A
authors: DA
authors: Bloch
authors: Marina
authors: M
authors: Verkhovskaya
mesh_terms: D020134:Catalytic Domain; D042967:Electron Transport Complex I / Q000737:chemistry* / Q000378:metabolism*; D004926:Escherichia coli / Q000737:chemistry / Q000201:enzymology*; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D005486:Flavin Mononucleotide / Q000378:metabolism; D018698:Glutamic Acid / Q000737:chemistry; D008958:Models, Molecular; D009154:Mutation; D009243:NAD / Q000378:metabolism; D010084:Oxidation-Reduction; D010100:Oxygen / Q000378:metabolism; D011485:Protein Binding; D011808:Quinone Reductases / Q000737:chemistry / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism*; D014451:Ubiquinone / Q000378:metabolism
chemical_list: D029968:Escherichia coli Proteins; D017382:Reactive Oxygen Species; D009243:NAD; D014451:Ubiquinone; D018698:Glutamic Acid; D005486:Flavin Mononucleotide; C502197:NuoF protein, E coli; D011808:Quinone Reductases; D042967:Electron Transport Complex I; D010100:Oxygen