title: A Comprehensive Analytical Strategy To Identify Malondialdehyde-Modified Proteins and Peptides.
abstract: Mass spectrometric-based proteomics is a powerful tool to analyze post-translationally modified proteins. Carbonylation modifications that result from oxidative lipid breakdown are a class of post-translational modifications that are poorly characterized with respect to protein targets and function. This is partly due to the lack of dedicated mass spectrometry-based technologies to facilitate the analysis of these modifications. Here, we present a comprehensive approach to identify malondialdehyde-modified proteins and peptides. Malondialdehyde is among the most abundant of the lipid peroxidation products; and malondialdehyde-derived adducts on proteins have been implicated in cardiovascular diseases, neurodegenerative disorders, and other clinical conditions. Our integrated approach targets three levels of the overall proteomic workflow: (i) sample preparation, by employing a targeted enrichment strategy; (ii) high-performance liquid chromatography, by using a gradient optimized for the separation of the modified peptides; and (iii) tandem mass spectrometry, by improving the spectral quality of very low-abundance peptides. By applying the optimized procedure to a whole cell lysate spiked with a low amount of malondialdehyde-modified proteins, we were able to identify up to 350 different modified peptides and localize the modification to a specific lysine residue. This methodology allows the comprehensive analysis of malondialdehyde-modified proteins.
authors.affiliation: CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences , 1090, Vienna, Austria.
authors.forename: Juliane
authors.identifier: 0000-0002-5579-4587
authors.initials: J
authors.lastname: Weißer
authors.affiliation: CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences , 1090, Vienna, Austria.
authors.forename: Claudia
authors.initials: C
authors.lastname: Ctortecka
authors.affiliation: CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences , 1090, Vienna, Austria.|Department of Laboratory Medicine, Medical University of Vienna , 1090, Vienna, Austria.
authors.forename: Clara J
authors.initials: CJ
authors.lastname: Busch
authors.affiliation: CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences , 1090, Vienna, Austria.|Department of Internal Medicine I, Medical University of Vienna , 1090, Vienna, Austria.
authors.forename: Shane R
authors.initials: SR
authors.lastname: Austin
authors.affiliation: Department of Internal Medicine I, Medical University of Vienna , 1090, Vienna, Austria.
authors.forename: Karin
authors.initials: K
authors.lastname: Nowikovsky
authors.affiliation: Graduate School of Bioagricultural Sciences, Nagoya University , Nagoya 464-8601, Japan.
authors.forename: Koji
authors.initials: K
authors.lastname: Uchida
authors.affiliation: CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences , 1090, Vienna, Austria.|Department of Laboratory Medicine, Medical University of Vienna , 1090, Vienna, Austria.
authors.forename: Christoph J
authors.initials: CJ
authors.lastname: Binder
authors.affiliation: CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences , 1090, Vienna, Austria.
authors.forename: Keiryn L
authors.initials: KL
authors.lastname: Bennett
mesh_terms: D008315:Malondialdehyde / Q000032:analysis*; D013058:Mass Spectrometry; D015394:Molecular Structure; D010455:Peptides / Q000737:chemistry*; D011506:Proteins / Q000737:chemistry*
chemical_list: D010455:Peptides; D011506:Proteins; D008315:Malondialdehyde