title: Proteomic analysis of 4-hydroxynonenal (4-HNE) modified proteins in liver mitochondria from chronic ethanol-fed rats.
abstract: Chronic ethanol-mediated oxidative stress and lipid peroxidation increases the levels of various reactive lipid species including 4-hydroxynonenal (4-HNE), which can subsequently modify proteins in the liver. It has been proposed that 4-HNE modification adversely affects the structure and/or function of mitochondrial proteins, thereby impairing mitochondrial metabolism. To determine whether chronic ethanol consumption increases levels of 4-HNE modified proteins in mitochondria, male rats were fed control and ethanol-containing diets for 5 weeks and mitochondrial samples were analyzed using complementary proteomic methods. Five protein bands (approx. 35, 45, 50, 70, and 90kDa) showed strong immunoreactivity for 4-HNE modified proteins in liver mitochondria from control and ethanol-fed rats when proteins were separated by standard 1D SDS-PAGE. Using high-resolution proteomic methods (2D IEF/SDS-PAGE and BN-PAGE) we identified several mitochondrial proteins immunoreactive for 4-HNE, which included mitofilin, dimethylglycine dehydrogenase, choline dehydrogenase, electron transfer flavoprotein α, cytochrome c1, enoyl CoA hydratase, and cytochrome c. The electron transfer flavoprotein α consistently showed increased 4-HNE immunoreactivity in mitochondria from ethanol-fed rats as compared to mitochondria from the control group. Increased 4-HNE reactivity was also detected for dimethylglycine dehydrogenase, enoyl CoA hydratase, and cytochrome c in ethanol samples when mitochondria were analyzed by BN-PAGE. In summary, this work identifies new targets of 4-HNE modification in mitochondria and provides useful information needed to better understand the molecular mechanisms underpinning chronic ethanol-induced mitochondrial dysfunction and liver injury.
authors.affiliation: Department of Medicine, Division of Nephrology, University of Alabama at Birmingham, Birmingham, AL, USA; Department of Environmental Health Sciences, University of Alabama at Birmingham, Birmingham, AL, USA.
authors.forename: Kelly K
authors.initials: KK
authors.lastname: Andringa
authors.affiliation: Department of Pathology, Division of Molecular and Cellular Pathology, University of Alabama at Birmingham, Birmingham, AL, USA.
authors.forename: Uduak S
authors.initials: US
authors.lastname: Udoh
authors.affiliation: Department of Pathology, Division of Molecular and Cellular Pathology, University of Alabama at Birmingham, Birmingham, AL, USA.
authors.forename: Aimee
authors.initials: A
authors.lastname: Landar
authors.affiliation: Department of Environmental Health Sciences, University of Alabama at Birmingham, Birmingham, AL, USA; Department of Pathology, Division of Molecular and Cellular Pathology, University of Alabama at Birmingham, Birmingham, AL, USA. Electronic address: sbailey@uab.edu.
authors.forename: Shannon M
authors.initials: SM
authors.lastname: Bailey
mesh_terms: D000447:Aldehydes / Q000378:metabolism*; D000818:Animals; D002908:Chronic Disease; D000431:Ethanol / Q000633:toxicity*; D008108:Liver Diseases, Alcoholic / Q000378:metabolism* / Q000473:pathology; D008297:Male; D008930:Mitochondria, Liver / Q000378:metabolism* / Q000473:pathology; D024101:Mitochondrial Proteins / Q000378:metabolism*; D011499:Protein Processing, Post-Translational / Q000187:drug effects*; D040901:Proteomics; D051381:Rats; D017207:Rats, Sprague-Dawley
chemical_list: D000447:Aldehydes; D024101:Mitochondrial Proteins; D000431:Ethanol; C027576:4-hydroxy-2-nonenal
keywords: 4-Hydroxynonenal; Alcohol; Liver; Mitochondria; Posttranslational modification; Proteome