Protein Modification by Endogenously Generated Lipid Electrophiles: Mitochondria as the Source and Target
William N Beavers1, Kristie L Rose1, James J Galligan1
1A.B. Hancock Memorial Laboratory for Cancer Research, Departments of Chemistry, ‡Biochemistry, §Pharmacology, ∥Biomedical Informatics, ⊥Vanderbilt Mass Spectrometry Research Center, Vanderbilt Institute for Chemical Biology, Vanderbilt Center in Molecular Toxicology, Vanderbilt Ingram Cancer Center, Vanderbilt University , Nashville, Tennessee 37232, United States.
Abstract:
Determining the impact of lipid electrophile-mediated protein damage that occurs during oxidative stress requires a comprehensive analysis of electrophile targets adducted under pathophysiological conditions. Incorporation of ω-alkynyl linoleic acid into the phospholipids of macrophages prior to activation by Kdo2-lipid A, followed by protein extraction, click chemistry, and streptavidin affinity capture, enabled a systems-level survey of proteins adducted by lipid electrophiles generated endogenously during the inflammatory response. Results revealed a dramatic enrichment for membrane and mitochondrial proteins as targets for adduction. A marked decrease in adduction in the presence of MitoTEMPO demonstrated a primary role for mitochondrial superoxide in electrophile generation and indicated an important role for mitochondria as both a source and target of lipid electrophiles, a finding that has not been revealed by prior studies using exogenously provided electrophiles.
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