Covalent modification of cytochrome c exposed to trans,trans-2,4-decadienal.
Modification of biomolecules by reactive aldehydes is believed to play a role in biological processes, including aging, atherosclerosis, and Alzheimer's disease. Here, the modification of cytochrome c promoted by trans, trans-2,4-decadienal (DDE) was investigated. Matrix-assisted laser desorption/ionization time-of-flight experiments indicated increases in the molecular weight of cytochrome c, consistent with the formation of DDE adducts. Our data show that the protein modification was time-, pH-, and DDE concentration-dependent, leading to the formation of at least six adducts after 2 h of incubation at pH 7.4. Electrospray ionization quantitative TOF mass spectrometry analysis of tryptic digests indicated that His-33, Lys-39, Lys-72, and Lys-100 were modified by DDE. These adducts could have significant effects considering that His-33, Lys-72, and Lys-100 are present in clusters of basic amino acid residues, which are believed to participate in the interaction of cytochrome c with cardiolipin in the inner mitochondrial membrane and cytochrome c oxidase. A blue shift in the cytochrome c Soret band from 409 to 406 nm was also observed after DDE reaction, indicating heme crevice opening and displacement of heme sixth ligand (Met-80) coordination in modified protein. The covalent modifications in cytochrome c could play a role in mitochondrial dysfunction associated with oxidative stress.
Departamento de Bioquímica, Instituto de Química, Universidade de São Paulo, Av. Prof. Lineu Prestes 748, CEP 05508-900, São Paulo, Brazil.
Carlos A O
CA
Sigolo
Paolo
P
Di Mascio
Marisa H G
MH
Medeiros
D000447:Aldehydes / Q000737:chemistry*; D000595:Amino Acid Sequence; D002308:Cardiolipins / Q000737:chemistry / Q000378:metabolism; D045304:Cytochromes c / Q000737:chemistry* / Q000378:metabolism; D003576:Electron Transport Complex IV / Q000378:metabolism; D006639:Histidine / Q000737:chemistry / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D008239:Lysine / Q000737:chemistry / Q000378:metabolism; D051336:Mitochondrial Membranes / Q000378:metabolism*; D008969:Molecular Sequence Data; D008970:Molecular Weight; D018384:Oxidative Stress / Q000187:drug effects* / Q000502:physiology; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013997:Time Factors
D000447:Aldehydes; D002308:Cardiolipins; C057349:2,4-decadienal; D006639:Histidine; D045304:Cytochromes c; D003576:Electron Transport Complex IV; D008239:Lysine