Protein targets for carbonylation by 4-hydroxy-2-nonenal in rat liver mitochondria

J Proteomics. 2011 Oct 19;74(11):2370-9. doi: 10.1016/j.jprot.2011.07.009. Epub 2011 Jul 23.


Protein carbonylation has been associated with various pathophysiological processes. A representative reactive carbonyl species (RCS), 4-hydroxy-2-nonenal (HNE), has been implicated specifically as a causative factor for the initiation and/or progression of various diseases. To date, however, little is known about the proteins and their modification sites susceptible to "carbonyl stress" by this RCS, especially in the liver. Using chemoprecipitation based on a solid-phase hydrazine chemistry coupled with LC-MS/MS bottom-up approach and database searching, we identified several protein-HNE adducts in isolated rat liver mitochondria upon HNE exposure. The identification of selected major protein targets, such as the ATP synthase β-subunit, was further confirmed by immunoblotting and a gel-based approach in combination with LC-MS/MS. A network was also created based on the identified protein targets, which showed that the main protein interactions were associated with cell death, tumor morphology and drug metabolism, implicating the toxic nature of HNE in the liver mitoproteome. The functional consequence of carbonylation was illustrated by its detrimental impact on the activity of ATP synthase, a representative major mitochondrial protein target for HNE modifications.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Aldehydes / pharmacology*
  • Amino Acid Sequence
  • Animals
  • Drug Delivery Systems
  • Liver / chemistry
  • Liver / drug effects
  • Liver / metabolism
  • Male
  • Metabolic Networks and Pathways
  • Mitochondria, Liver / chemistry
  • Mitochondria, Liver / drug effects*
  • Mitochondria, Liver / metabolism
  • Mitochondrial Proteins / chemistry
  • Mitochondrial Proteins / drug effects
  • Mitochondrial Proteins / isolation & purification
  • Mitochondrial Proteins / metabolism*
  • Mitochondrial Proton-Translocating ATPases / chemistry
  • Mitochondrial Proton-Translocating ATPases / metabolism
  • Peptide Fragments / chemistry
  • Peptide Fragments / metabolism
  • Protein Carbonylation / drug effects*
  • Protein Carbonylation / physiology
  • Rats
  • Rats, Sprague-Dawley
  • Reactive Oxygen Species / chemistry
  • Reactive Oxygen Species / metabolism


  • Aldehydes
  • Mitochondrial Proteins
  • Peptide Fragments
  • Reactive Oxygen Species
  • Mitochondrial Proton-Translocating ATPases
  • 4-hydroxy-2-nonenal