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A single amino acid residue controls ROS production in the respiratory C omplex I from E scherichia coli
Author(s) -
Knuuti Juho,
Belevich Galina,
Sharma Vivek,
Bloch Dmitry A.,
Verkhovskaya Marina
Publication year - 2013
Publication title -
molecular microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.857
H-Index - 247
eISSN - 1365-2958
pISSN - 0950-382X
DOI - 10.1111/mmi.12424
Subject(s) - flavin mononucleotide , biology , population , escherichia coli , biochemistry , flavin group , nad+ kinase , nicotinamide adenine dinucleotide , enzyme , flavin adenine dinucleotide , redox , flavoprotein , nicotinamide mononucleotide , stereochemistry , cofactor , chemistry , demography , organic chemistry , sociology , gene
Summary Reactive oxygen species ( ROS ) production by respiratory C omplex I from E scherichia 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 C omplex I towards dioxygen ( O 2 ). 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 C omplex I . The analysis of the kinetic model of ROS production allowed us to propose that the population of C omplex I with reduced FMN is always low in the wild‐type enzyme even at low ambient redox potentials, minimizing the rate of reaction with O 2 in contrast to E95Q variant.