title: Mass spectrometry-based quantification of myocardial protein adducts with acrolein in an in vivo model of oxidative stress.
abstract: Acrolein (ACR) exposure leads to the formation of protein-ACR adducts. Protein modification by ACR has been associated with various chronic diseases including cardiovascular and neurodegenerative diseases. Here, we report an analytical strategy that enables the quantification of Michael-type protein adducts of ACR in mitochondrial proteome samples using liquid chromatography in combination with tandem mass spectrometry and selected ion monitoring (LC-MS/MS SRM) analysis. Our approach combines site-specific identification and relative quantification at the peptide level of protein-ACR adducts in relation to the unmodified protein thiol pool. Treatment of 3-month-old rats with CCl(4) , an established in vivo model of acute oxidative stress, resulted in significant increases in the ratios of distinct ACR-adducted peptides to the corresponding unmodified thiol-peptides obtained from proteins that were isolated from cardiac mitochondria. The mitochondrial proteins that were found adducted by ACR were malate dehydrogenase, NADH dehydrogenase [ubiquinone] flavoprotein 1, cytochrome c oxidase subunit VIb isoform 1, ATP synthase d chain, and ADP/ATP translocase 1. The findings indicate that protein modification by ACR has potential value as an index of mitochondrial oxidative stress.
authors.affiliation: Department of Chemistry, Oregon State University, Corvallis OR 97330, USA.
authors.forename: Jianyong
authors.initials: J
authors.lastname: Wu
authors.forename: Jan F
authors.initials: JF
authors.lastname: Stevens
authors.forename: Claudia S
authors.initials: CS
authors.lastname: Maier
mesh_terms: D000171:Acrolein / Q000032:analysis* / Q000737:chemistry; D000595:Amino Acid Sequence; D000818:Animals; D002725:Chloroform / Q000633:toxicity; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D004195:Disease Models, Animal; D008291:Malate Dehydrogenase / Q000737:chemistry / Q000378:metabolism; D008297:Male; D008929:Mitochondria, Heart / Q000378:metabolism; D000226:Mitochondrial ADP, ATP Translocases / Q000737:chemistry / Q000378:metabolism; D024101:Mitochondrial Proteins / Q000032:analysis* / Q000737:chemistry / Q000378:metabolism; D008969:Molecular Sequence Data; D009206:Myocardium / Q000378:metabolism*; D018384:Oxidative Stress; D010455:Peptides / Q000737:chemistry / Q000378:metabolism; D020543:Proteome; D051381:Rats; D011916:Rats, Inbred F344; D053719:Tandem Mass Spectrometry / Q000379:methods*
chemical_list: D024101:Mitochondrial Proteins; D010455:Peptides; D020543:Proteome; D000171:Acrolein; D002725:Chloroform; D000226:Mitochondrial ADP, ATP Translocases; D008291:Malate Dehydrogenase