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Cytochrome c-promoted cardiolipin oxidation generates singlet molecular oxygen

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Abstract

The interaction of cytochrome c (cyt c) with cardiolipin (CL) induces protein conformational changes that favor peroxidase activity. This process has been correlated with CL oxidation and the induction of cell death. Here we report evidence demonstrating the generation of singlet molecular oxygen [O2(1Δg)] by a cyt c—CL complex in a model membrane containing CL. The formation of singlet oxygen was directly evidenced by luminescence measurements at 1270 nm and by chemical trapping experiments. Singlet oxygen generation required cyt c—CL binding and occurred at pH values higher than 6, consistent with lipid—protein interactions involving fully deprotonated CL species and positively charged residues in the protein. Moreover, singlet oxygen formation was specifically observed for tetralinoleoyl CL species and was not observed with monounsaturated and saturated CL species. Our results show that there are at least two mechanisms leading to singlet oxygen formation: one with fast kinetics involving the generation of singlet oxygen directly from CL hydroperoxide decomposition and the other involving CL oxidation. The contribution of the first mechanism was clearly evidenced by the detection of labeled singlet oxygen [18O2(1Δg)] from liposomes supplemented with 18-oxygen-labeled CL hydroperoxides. However quantitative analysis showed that singlet oxygen yield from CL hydroperoxides was minor (<5%) and that most of the singlet oxygen is formed from the second mechanism. Based on these data and previous findings we propose a mechanism of singlet oxygen generation through reactions involving peroxyl radicals (Russell mechanism) and excited triplet carbonyl intermediates (energy transfer mechanism).

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Authors and Affiliations

  1. Departamento de Bioquímica, Instituto de Química, Universidade de São Paulo, CP26077, CEP 05513-970, São Paulo, SP, Brazil

    Sayuri Miyamoto, Daniela Cunha, Graziella E. Ronsein, Marisa H. G. Medeiros & Paolo Di Mascio

  2. Centro de Ciências Naturais e Humanas, Universidade Federal do ABC, Rua Santa Adélia 166, Bairro Bangu, CEP 09210-170, Santo André, SP, Brazil

    Iseli L. Nantes & Priscila A. Faria

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  1. Sayuri Miyamoto
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  2. Iseli L. Nantes
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  3. Priscila A. Faria
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  4. Daniela Cunha
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  5. Graziella E. Ronsein
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  6. Marisa H. G. Medeiros
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Correspondence to Sayuri Miyamoto or Paolo Di Mascio.

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Electronic supplementary information (ESI) available: Analysis of CL hydroperoxide content; MS analysis of 18-oxygen labeled CL hydroperoxides; DHPNO2 emission signal; and visible light emission enhancement by DBAS. See DOI: 10.1039/c2pp25119a

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Miyamoto, S., Nantes, I.L., Faria, P.A. et al. Cytochrome c-promoted cardiolipin oxidation generates singlet molecular oxygen. Photochem Photobiol Sci 11, 1536–1546 (2012). https://doi.org/10.1039/c2pp25119a

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