title: Cytochrome c adducts with PCB quinoid metabolites.
abstract: Polychlorinated biphenyls (PCBs) are a group of 209 individual congeners widely used as industrial chemicals. PCBs are found as by-products in dye and paint manufacture and are legacy, ubiquitous, and persistent as human and environmental contaminants. PCBs with fewer chlorine atoms may be metabolized to hydroxy- and dihydroxy-metabolites and further oxidized to quinoid metabolites both in vitro and in vivo. Specifically, quinoid metabolites may form adducts on nucleophilic sites within cells. We hypothesized that the PCB-quinones covalently bind to cytochrome c and, thereby, cause defects in the function of cytochrome c. In this study, synthetic PCB quinones, 2-(4'-chlorophenyl)-1,4-benzoquinone (PCB3-pQ), 4-4'-chlorophenyl)-1,2-benzoquinone (PCB3-oQ), 2-(3', 5'-dichlorophenyl)-1,4-benzoquinone, 2-(3',4', 5'-trichlorophenyl)-1,4-benzoquinone, and 2-(4'-chlorophenyl)-3,6-dichloro-1,4-benzoquinone, were incubated with cytochrome c, and adducts were detected by liquid chromatography-mass spectrometry (LC-MS) and matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI TOF). Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) was employed to separate the adducted proteins, while trypsin digestion and liquid chromatography-tandem mass spectrometry (LC-MS/MS) were applied to identify the amino acid binding sites on cytochrome c. Conformation change of cytochrome c after binding with PCB3-pQ was investigated by SYBYL-X simulation and cytochrome c function was examined. We found that more than one molecule of PCB-quinone may bind to one molecule of cytochrome c. Lysine and glutamic acid were identified as the predominant binding sites. Software simulation showed conformation changes of adducted cytochrome c. Additionally, cross-linking of cytochrome c was observed on the SDS-PAGE gel. Cytochrome c was found to lose its function as electron acceptor after incubation with PCB quinones. These data provide evidence that the covalent binding of PCB quinone metabolites to cytochrome c may be included among the toxic effects of PCBs.
authors.affiliation: Interdisciplinary Graduate Program in Human Toxicology, Graduate College, The University of Iowa, Iowa City, IA, USA.|Department of Occupational & Environmental Health, College of Public Health, The University of Iowa, 100 Oakdale Campus 214 IREH, Iowa City, IA, 52242-5000, USA.
authors.forename: Miao
authors.initials: M
authors.lastname: Li
authors.affiliation: High Resolution Mass Spectrometry Facility, The University of Iowa, Iowa City, IA, USA.
authors.forename: Lynn M
authors.initials: LM
authors.lastname: Teesch
authors.affiliation: Interdisciplinary Graduate Program in Human Toxicology, Graduate College, The University of Iowa, Iowa City, IA, USA.|College of Pharmacy, The University of Iowa, Iowa City, IA, USA.
authors.forename: Daryl J
authors.initials: DJ
authors.lastname: Murry
authors.affiliation: Proteomics Facility, The University of Iowa, Iowa City, IA, USA.
authors.forename: R Marshal
authors.initials: RM
authors.lastname: Pope
authors.affiliation: Proteomics Facility, The University of Iowa, Iowa City, IA, USA.
authors.forename: Yalan
authors.initials: Y
authors.lastname: Li
authors.affiliation: Interdisciplinary Graduate Program in Human Toxicology, Graduate College, The University of Iowa, Iowa City, IA, USA.|Department of Occupational & Environmental Health, College of Public Health, The University of Iowa, 100 Oakdale Campus 214 IREH, Iowa City, IA, 52242-5000, USA.
authors.forename: Larry W
authors.initials: LW
authors.lastname: Robertson
authors.affiliation: Interdisciplinary Graduate Program in Human Toxicology, Graduate College, The University of Iowa, Iowa City, IA, USA. gabriele-ludewig@uiowa.edu.|Department of Occupational & Environmental Health, College of Public Health, The University of Iowa, 100 Oakdale Campus 214 IREH, Iowa City, IA, 52242-5000, USA. gabriele-ludewig@uiowa.edu.
authors.forename: Gabriele
authors.initials: G
authors.lastname: Ludewig
mesh_terms: D002853:Chromatography, Liquid; D045304:Cytochromes c / Q000737:chemistry* / Q000378:metabolism; D006801:Humans; D015394:Molecular Structure; D010084:Oxidation-Reduction; D011078:Polychlorinated Biphenyls / Q000737:chemistry* / Q000378:metabolism; D053719:Tandem Mass Spectrometry
chemical_list: D045304:Cytochromes c; D011078:Polychlorinated Biphenyls
keywords: 4-Chlorobiphenyl; Cytochrome c; Post-translational modification; Protein adducts; Protein cross-linking; Quinoid metabolites