title: Cytochrome c reacts with cholesterol hydroperoxides to produce lipid- and protein-derived radicals.
abstract: Lipid peroxidation is a well-known process that has been implicated in many diseases. Recent evidence has shown that mitochondrial cholesterol levels are increased under specific conditions, making it an important target for peroxidation inside the mitochondria. Cholesterol peroxidation generates, as primary products, several hydroperoxides (ChOOH), which can react with transition metals and metalloproteins. In this sense, cytochrome c (CYTC), a heme protein largely found in the mitochondria, becomes a candidate to react with ChOOH. Using CYTC associated with SDS micelles to mimic mitochondrial conditions, we show that ChOOH induces dose-dependent CYTC Soret band bleaching, indicating that it is using ChOOH as a substrate. This reaction leads to protein oligomerization, suggesting the formation of a protein radical that, subsequently, recombines, giving dimers, trimers, and tetramers. EPR experiments confirmed the production of carbon-centered radicals from both protein and lipid in the presence of ChOOH. Similar results were obtained with linoleic acid hydroperoxides (LAOOH). In addition, replacing SDS micelles by cardiolipin-containing liposomes as the mitochondrial mimetic led to similar results with either ChOOH or LAOOH. Importantly, kinetic experiments show that CYTC bleaching is faster with ChOOH than with H2O2, suggesting that these hydroperoxides could be relevant substrates for CYTC peroxidase-like activity in biological media. Altogether, these results show that CYTC induces homolytic cleavage of lipid-derived hydroperoxides, producing lipid and protein radicals.
authors: †Departamento de Bioquímica, Instituto de Química, Universidade de São Paulo, São Paulo, SP 05508-000, Brazil.
authors: Thiago C
authors: TC
authors: Genaro-Mattos
authors: †Departamento de Bioquímica, Instituto de Química, Universidade de São Paulo, São Paulo, SP 05508-000, Brazil.|‡Departamento de Química e Exatas, Universidade Estadual do Sudoeste da Bahia, Jequié, BA 45200-000, Brazil.
authors: Raphael F
authors: RF
authors: Queiroz
authors: †Departamento de Bioquímica, Instituto de Química, Universidade de São Paulo, São Paulo, SP 05508-000, Brazil.
authors: Daniela
authors: D
authors: Cunha
authors: †Departamento de Bioquímica, Instituto de Química, Universidade de São Paulo, São Paulo, SP 05508-000, Brazil.
authors: Patricia P
authors: PP
authors: Appolinario
authors: †Departamento de Bioquímica, Instituto de Química, Universidade de São Paulo, São Paulo, SP 05508-000, Brazil.
authors: Paolo
authors: P
authors: Di Mascio
authors: §Centro de Ciências Naturais e Humanas, Universidade Federal do ABC, Santo André, SP 09210-580, Brazil.
authors: Iseli L
authors: IL
authors: Nantes
authors: †Departamento de Bioquímica, Instituto de Química, Universidade de São Paulo, São Paulo, SP 05508-000, Brazil.
authors: Ohara
authors: O
authors: Augusto
authors: †Departamento de Bioquímica, Instituto de Química, Universidade de São Paulo, São Paulo, SP 05508-000, Brazil.
authors: Sayuri
authors: S
authors: Miyamoto
mesh_terms: D000818:Animals; D002417:Cattle; D002784:Cholesterol / Q000031:analogs & derivatives* / Q000737:chemistry; D045304:Cytochromes c / Q000737:chemistry*; D005609:Free Radicals / Q000737:chemistry*; D006861:Hydrogen Peroxide / Q000737:chemistry; D007700:Kinetics; D008041:Linoleic Acids / Q000737:chemistry; D015227:Lipid Peroxidation; D008054:Lipid Peroxides / Q000737:chemistry; D008081:Liposomes; D008823:Micelles; D058105:Polymerization; D011725:Pyridines / Q000737:chemistry; D012967:Sodium Dodecyl Sulfate
chemical_list: C450079:(pyridyl-4-N-oxide)-N-tert-butylnitrone; D005609:Free Radicals; D008041:Linoleic Acids; D008054:Lipid Peroxides; D008081:Liposomes; D008823:Micelles; D011725:Pyridines; C008285:linoleic acid hydroperoxide; D012967:Sodium Dodecyl Sulfate; C002843:cholesterol hydroperoxide; D045304:Cytochromes c; D002784:Cholesterol; D006861:Hydrogen Peroxide