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Peroxidase complexes of cytochrome c with anionic lipids: structural pre-requisites, mechanisms, and cytotoxic effects.

Vlasova-II; Kapralov-AA; Jiang-J; Basova-LV; Belikova-NA; Tyurin-VA; Tyurina-YY; Martin-J; Glumac-A; Bayir-H; Kagan-VE
Toxicologist 2007 Mar; 96(1):61
Interaction of cytochrome c (cyt c) with a mitochondria-specific cardiolipin (CL) confers peroxidase activity on the protein resulting in selective CL oxidation and release of proapoptotic factors. Because the complex cyt c/CL is stabilized by a combination of electrostatic and hydrophobic interactions, we determined the extent to which other anionic lipids - phosphatidic acid (PA), phosphatidylserine (PS), phosphatidylinositolphosphates (PIP), and phosphatidylcholine (PC) as a control - are effective in inducing the peroxidase activity of cyt c. EPR spectroscopy of nitrosylated cyt c, optical spectroscopy and measurements of tryptophan fluorescence demonstrated that cyt c interaction with anionic lipids induced protein unfolding accompanied by an exchange and loss of axial ligands of heme iron so that cyt c heme became more accessible for the interaction with small molecules like NO or H2O2. Using several peroxidase substrates we showed that all anionic lipids activated peroxidase activity of cyt c in a dose-dependent manner with the efficiency decreasing in the row: CL=PA>PIP2>PIP3>PS. Recombination of protein-derived radicals formed in peroxidase reaction caused oligomerization of cyt c and formation of protein-lipid aggregates detectable by PAGE and Western blotting. In line with this, Western blotting revealed the formation of cyt c aggregates after its incubation in the presence of H2O2 with membrane (but not cytosolic S-100) fraction of brain homogenates. Oxidation products of anionic lipids were detected after induction of apoptosis in HeLa cells and mouse embryonic wild type cells but not in cyt c-deficient cells. Overall, our study identified anionic lipids as physiologically relevant regulators of peroxidase activity of cyt c in mitochondria and other cell compartments, particularly during apoptosis.
Cell-alteration; Cell-damage; Cellular-reactions; Chemical-analysis; Chemical-composition; Chemical-indicators; Chemical-synthesis; Biomarkers; Animal-studies; Physiological-chemistry; Physiological-response; Physiological-effects
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The Toxicologist. Society of Toxicology 46th Annual Meeting and ToxExpo, March 25-29, 2007, Charlotte, North Carolina
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University of Pittsburgh at Pittsburgh