Not Only Oxidation of Cardiolipin Affects the Affinity of Cytochrome c for Lipid Bilayers

• Published in: The journal of physical chemistry. B 2014-10, Vol.118 (41), p.11863-11872
• Main Authors: Kawai, Cintia, Ferreira, Juliana C, Baptista, Mauricio S, Nantes, Iseli L
• Format: Article
• Language: English
• Subjects: Affinity; Animals; Binding; Cardiolipins - chemistry; Circular Dichroism; Circularity; Constants; Cytochromes; Cytochromes c - chemistry; Dichroism; Fish Proteins - chemistry; Fluorescence; Horses; Hydrophobic and Hydrophilic Interactions; Lipid Bilayers - chemistry; Lipids; Liposomes - chemistry; Mitochondrial Membranes - chemistry; Models, Biological; Molecular Structure; Myocardium - chemistry; Oxidation-Reduction; Phosphatidylcholines - chemistry; Phosphatidylethanolamines - chemistry; Phosphorylcholine - analogs & derivatives; Phosphorylcholine - chemistry; Pyrenes - chemistry; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; Tuna
• ISSN: 1520-6106; 1520-5207
• DOI: 10.1021/jp504518g

Fluorescence quenching of lipid-bound pyrene was used to assess the binding of cytochrome c (cyt c) to liposomes that mimic the inner mitochondrial membrane (IMM) POPC/​DOPE/​TOCL, with the conditions that it did or did not contain oxidized phosphatidyl­choline molecules, i.e., 1-O-hexadecyl-2-azelaoyl-sn-glycero-3-phospho­choline (PazePC), or a mixture of two hydro­peroxide isomers derived from POPC (POPCOX). The binding isotherms reveal two dissociation constants, K D 1 and K D 2, representing, respectively, the low- and high-affinity states of the membrane. These dissociation constants probably are due to the lipid reorganization promoted by cyt c, as observed in giant unilamellar vesicles that contain fluorescent cardiolipin (CL). The presence of PazePC, which has a nonreactive carboxylic group, increased the K D 1 and K D 2 values 1.2- and 4.5-fold, respectively. The presence of POPCOX which has a reactive peroxide group, decreased the K D 1 value 1.5-fold, increased the K D 2 value 10-fold, and significantly reduced the salt-induced detachment of cyt c. MALDI-TOF spectrometry analysis of cyt c incubated with liposomes containing POPCox demonstrated a mass increase corresponding to the formation of nonenal adducts as hydrophobic anchors. Electronic absorption spectroscopy, circular dichroism, and magnetic circular dichroism demonstrated that all of the lipids studied promoted changes in the cyt c coordination sphere. Therefore, in the presence of CL, the oxidation of zwitterionic lipids also promotes changes in the cyt c structure and in the affinity for lipid bilayers.