Global analysis of protein damage by the lipid electrophile 4-hydroxy-2-nonenal

Simona G. Codreanu, Bing Zhang, Scott M. Sobecki, David D Billheimer, Daniel C. Liebler

Research output: Contribution to journalArticle

108 Citations (Scopus)

Abstract

Lipid peroxidation yields a variety of electrophiles, which are thought to contribute to the molecular pathogenesis of diseases involving oxidative stress, yet little is known of the scope of protein damage caused by lipid electrophiles. We identified protein targets of the prototypical lipid electrophile 4-hydroxy-2-nonenal (HNE) in RKO cells treated with 50 or 100 μM HNE. HNE Michael adducts were biotinylated by reaction with biotinamidohexanoic acid hydrazide, captured with streptavidin, and the captured proteins were resolved by one dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis, digested with trypsin, and identified by liquid chromatography-tandem mass spectrometry. Of the 1500+ proteins identified, 417 displayed a statistically significant increase in adduction with increasing HNE exposure concentration. We further identified 18 biotin hydrazide-modified, HNE-adducted peptides by specific capture using anti-biotin antibody and analysis by high resolution liquid chromatography-tandem mass spectrometry. A subset of the identified HNE targets were validated with a streptavidin capture and immunoblotting approach, which enabled detection of adducts at HNE exposures as low as 1 μM. Protein interaction network analysis indicated several subsystems impacted by endogenous electrophiles in oxidative stress, including the 26 S proteasomal and chaperonin containing TCP-1 (CCT) systems involved in protein-folding and degradation, as well as the COP9 signalosome, translation initiation complex, and a large network of ribonucleoproteins. Global analyses of protein lipid electrophile adducts provide a systems-level perspective on the mechanisms of diseases involving oxidative stress.

Original languageEnglish (US)
Pages (from-to)670-680
Number of pages11
JournalMolecular and Cellular Proteomics
Volume8
Issue number4
DOIs
StatePublished - Apr 2009
Externally publishedYes

Fingerprint

Lipids
Oxidative stress
Proteins
Oxidative Stress
Streptavidin
Liquid chromatography
Tandem Mass Spectrometry
Liquid Chromatography
Mass spectrometry
Chaperonin Containing TCP-1
Protein Interaction Maps
Protein folding
Ribonucleoproteins
Protein Folding
Biotin
Electric network analysis
4-hydroxy-2-nonenal
Electrophoresis
Immunoblotting
Sodium Dodecyl Sulfate

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Analytical Chemistry
  • Medicine(all)

Cite this

Global analysis of protein damage by the lipid electrophile 4-hydroxy-2-nonenal. / Codreanu, Simona G.; Zhang, Bing; Sobecki, Scott M.; Billheimer, David D; Liebler, Daniel C.

In: Molecular and Cellular Proteomics, Vol. 8, No. 4, 04.2009, p. 670-680.

Research output: Contribution to journalArticle

Codreanu, Simona G. ; Zhang, Bing ; Sobecki, Scott M. ; Billheimer, David D ; Liebler, Daniel C. / Global analysis of protein damage by the lipid electrophile 4-hydroxy-2-nonenal. In: Molecular and Cellular Proteomics. 2009 ; Vol. 8, No. 4. pp. 670-680.
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