title: Peroxidative permeabilization of liposomes induced by cytochrome c/cardiolipin complex.
abstract: Interaction of cytochrome c with mitochondrial cardiolipin converting this electron transfer protein into peroxidase is accepted to play an essential role in apoptosis. Cytochrome c/cardiolipin peroxidase activity was found here to cause leakage of carboxyfluorescein, sulforhodamine B and 3-kDa (but not 10-kDa) fluorescent dextran from liposomes. A marked decrease in the amplitude of the autocorrelation function was detected with a fluorescence correlation spectroscopy setup upon incubation of dye-loaded cardiolipin-containing liposomes with cytochrome c and H2O2, thereby showing release of fluorescent markers from liposomes. The cytochrome c/H2O2-induced liposome leakage was suppressed upon increasing the ionic strength, in contrast to the leakage provoked by Fe/ascorbate, suggesting that the binding of cyt c to negatively-charged membranes was required for the permeabilization process. The cyt c/H2O2-induced liposome leakage was abolished by cyanide presumably competing with H2O2 for coordination with the central iron atom of the heme in cyt c. The cytochrome c/H2O2 permeabilization activity was substantially diminished by antioxidants (trolox, butylhydroxytoluene and quercetin) and was precluded if fully saturated tetramyristoyl-cardiolipin was substituted for bovine heart cardiolipin. These data favor the involvement of oxidized cardiolipin molecules in membrane permeabilization resulting from cytochrome c/cardiolipin peroxidase activity. In agreement with previous observations, high concentrations of cyt c induced liposome leakage in the absence of H2O2, however this process was not sensitive to antioxidants and cyanide suggesting direct membrane poration by the protein without the involvement of lipid peroxidation.
authors.affiliation: Belozersky Institute of Physico-Chemical Biology, Lomonosov Moscow State University, Moscow 119991, Russia; Department of Bioengineering and Bioinformatics, Lomonosov Moscow State University, Moscow 119991, Russia.
authors.forename: Alexander M
authors.initials: AM
authors.lastname: Firsov
authors.affiliation: Belozersky Institute of Physico-Chemical Biology, Lomonosov Moscow State University, Moscow 119991, Russia. Electronic address: kotova@genebee.msu.ru.
authors.forename: Elena A
authors.initials: EA
authors.lastname: Kotova
authors.affiliation: Department of Medical Biophysics, N.I. Pirogov Russian National Research Medical University, Moscow, Russia.
authors.forename: Evgeniya A
authors.initials: EA
authors.lastname: Korepanova
authors.affiliation: Department of Medical Biophysics, N.I. Pirogov Russian National Research Medical University, Moscow, Russia.
authors.forename: Anatoly N
authors.initials: AN
authors.lastname: Osipov
authors.affiliation: Belozersky Institute of Physico-Chemical Biology, Lomonosov Moscow State University, Moscow 119991, Russia. Electronic address: antonen@genebee.msu.ru.
authors.forename: Yuri N
authors.initials: YN
authors.lastname: Antonenko
mesh_terms: D000465:Algorithms; D000818:Animals; D000975:Antioxidants / Q000494:pharmacology; D002084:Butylated Hydroxytoluene / Q000494:pharmacology; D002308:Cardiolipins / Q000737:chemistry* / Q000378:metabolism / Q000494:pharmacology; D002839:Chromans / Q000494:pharmacology; D045304:Cytochromes c / Q000737:chemistry* / Q000378:metabolism / Q000494:pharmacology; D003911:Dextrans / Q000737:chemistry / Q000378:metabolism; D005452:Fluoresceins / Q000737:chemistry / Q000378:metabolism; D006861:Hydrogen Peroxide / Q000494:pharmacology; D015227:Lipid Peroxidation / Q000187:drug effects; D008081:Liposomes / Q000737:chemistry* / Q000378:metabolism; D008956:Models, Chemical; D008958:Models, Molecular; D016877:Oxidants / Q000494:pharmacology; D010539:Permeability / Q000187:drug effects; D011485:Protein Binding / Q000187:drug effects; D011794:Quercetin / Q000494:pharmacology; D012235:Rhodamines / Q000737:chemistry / Q000378:metabolism; D013050:Spectrometry, Fluorescence
chemical_list: D000975:Antioxidants; D002308:Cardiolipins; D002839:Chromans; D003911:Dextrans; D005452:Fluoresceins; D008081:Liposomes; D016877:Oxidants; D012235:Rhodamines; D002084:Butylated Hydroxytoluene; C022027:lissamine rhodamine B; C024098:6-carboxyfluorescein; D045304:Cytochromes c; D011794:Quercetin; D006861:Hydrogen Peroxide; C010643:6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid
keywords: Antioxidant; Cardiolipin; Cytochrome c; Fluorescence correlation spectroscopy; Liposome leakage; Peroxidase