A single amino acid residue controls ROS production in the respiratory Complex I from Escherichia coli

作者：Knuuti Juho; Belevich Galina; Sharma Vivek; Bloch Dmitry A^*; Verkhovskaya Marina

来源：Molecular Microbiology, 2013, 90(6): 1190-1200.

DOI：10.1111/mmi.12424

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 (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 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 O-2 in contrast to E95Q variant.

• 出版日期2013-12