MccJ25 induce the superoxide and ROS overproduction in isolated mitochondria

V NIKLISON, MARIA; BEATRIZ F DE ARCURI; AUGUSTO BELLOMIO; CARLOS MINAHK; RD MORERO

Montevideo, Uruguay

Congreso; V Meeting of SFRBM Group and V Internacional conference of Peroxynitrite and Reactive Nitrogen Species.; 2007

SFRBM Group

Microcin J25 (MccJ25), a lasso peptide antibiotic  of 21 amino acid produced by an Escherichia coli strain, is active against close-related bacteria including certain human pathogens as some Salmonella and Shigella strains. MccJ25 act on the target cells inhibiting the RNAP activity, and operating on the cell membrane throughout the increasing of ROS concentration. The peptide displayed antimitochondrial activity dissipating the proton motive force with the concomitant inhibition of the ATP synthesis. In addition it inhibits the enzymatic activity of the cytochrome c reductase (complex III) of the respiratory chain. In this communication we studied the effect of MccJ25 on the reactive oxygen species (ROS) production. To overcome this point we measured the superoxide radical generation in sub mitochondrial particles (SMP), employing the method based on the ability of superoxide radical to reduce the partially acetylated cytochrome c. We determined ROS overproduction in intact isolated mitochondria evaluating the fluorescence of H2DCFDA-SE probe and measuring pyrimidin nucleotide oxidation through the NADPH fluorescence (lex 340 nm, lem 450 nm). The obtained results indicate that MccJ25 induce overproduction of superoxide radical in the SMP and ROS in the intact isolated mitochondria. Additionally, an increased oxidation of pyrimidin nucleotides was also observed. We hypothesizes that ROS overproduction would perturb the mitochondrial membrane which trigger a succession of event including the uptake of Calcium, and the consecutive activation of calcium uniport  and the permeability transition pore (MPT).