Oxidative modification of hepatic mitochondria protein thiols: effect of chronic alcohol consumption.

TitleOxidative modification of hepatic mitochondria protein thiols: effect of chronic alcohol consumption.
Publication TypeJournal Article
Year of Publication2004
AuthorsVenkatraman, A, Landar, A, Davis, AJ, Ulasova, E, Page, G, Murphy, MP, Darley-Usmar, V, Bailey, SM
JournalAm J Physiol Gastrointest Liver Physiol
Volume286
Issue4
PaginationG521-7
Date Published2004 Apr
ISSN0193-1857
KeywordsAldehyde Dehydrogenase, Animals, Central Nervous System Depressants, Chronic Disease, Electrophoresis, Polyacrylamide Gel, Energy Metabolism, Ethanol, Image Processing, Computer-Assisted, Immunoblotting, In Vitro Techniques, Male, Mitochondria, Liver, Molecular Weight, Oxidation-Reduction, Proteins, Rats, Rats, Sprague-Dawley, Reactive Nitrogen Species, Reactive Oxygen Species, Sulfhydryl Compounds
Abstract

Redox modification of mitochondrial proteins is thought to play a key role in regulating cellular function, although direct evidence to support this hypothesis is limited. Using an in vivo model of mitochondrial redox stress, ethanol hepatotoxicity, the modification of mitochondrial protein thiols was examined using a proteomics approach. Specific labeling of reduced thiols in the mitochondrion from the livers of control and ethanol-fed rats was achieved by using the thiol reactive compound (4-iodobutyl)triphenylphosphonium (IBTP). This molecule selectively accumulates in the organelle and can be used to identify thiol-containing proteins. Mitochondrial proteins that have been modified are identified by decreased labeling with IBTP using two-dimensional SDS-PAGE followed by immunoblotting with an antibody directed against the triphenylphosphonium moiety of the IBTP molecule. Analyses of these data showed a significant decrease in IBTP labeling of thiols present in specific mitochondria matrix proteins from ethanol-fed rats compared with their corresponding controls. These proteins were identified as the low-K(m) aldehyde dehydrogenase and glucose-regulated protein 78. The decrease in IBTP labeling in aldehyde dehydrogenase was accompanied by a decrease in specific activity of the enzyme. These data demonstrate that mitochondrial protein thiol modification is associated with chronic alcohol intake and might contribute to the pathophysiology associated with hepatic injury. Taken together, we have developed a protocol to chemically tag and select thiol-modified proteins that will greatly enhance efforts to establish posttranslational redox modification of mitochondrial protein in in vivo models of oxidative or nitrosative stress.

DOI10.1152/ajpgi.00399.2003
Alternate JournalAm. J. Physiol. Gastrointest. Liver Physiol.
Citation Key10.1152/ajpgi.00399.2003
PubMed ID14670822
Grant ListAA-13395 / AA / NIAAA NIH HHS / United States
AA-13682 / AA / NIAAA NIH HHS / United States
P30 CA-13148 / CA / NCI NIH HHS / United States
S10 RR-11329 / RR / NCRR NIH HHS / United States