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

Codreanu SG, Zhang B, Sobecki SM, Billheimer DD, Liebler DC - Mol. Cell Proteomics (2008)

Bottom Line: We identified protein targets of the prototypical lipid electrophile 4-hydroxy-2-nonenal (HNE) in RKO cells treated with 50 or 100 mum HNE.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.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry, Vanderbilt University School of Medicine, Nashville, Tennessee 37232, USA.

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 mum 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 mum. 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.

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Protein communities identified from the network of HNE-adducted proteins.
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f3: Protein communities identified from the network of HNE-adducted proteins.

Mentions: As HNE tends to coordinately target multiple proteins of a community, we used the software CFinder to explore the community structure in the network of HNE-targeted proteins. Fig. 3 depicts the network of protein communities identified by this analysis. Protein communities targeted by HNE included ribonucleoprotein complexes, translation initiation factors, proteasome, CCT complex, and COP signalosome complex.


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

Codreanu SG, Zhang B, Sobecki SM, Billheimer DD, Liebler DC - Mol. Cell Proteomics (2008)

Protein communities identified from the network of HNE-adducted proteins.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2667350&req=5

f3: Protein communities identified from the network of HNE-adducted proteins.
Mentions: As HNE tends to coordinately target multiple proteins of a community, we used the software CFinder to explore the community structure in the network of HNE-targeted proteins. Fig. 3 depicts the network of protein communities identified by this analysis. Protein communities targeted by HNE included ribonucleoprotein complexes, translation initiation factors, proteasome, CCT complex, and COP signalosome complex.

Bottom Line: We identified protein targets of the prototypical lipid electrophile 4-hydroxy-2-nonenal (HNE) in RKO cells treated with 50 or 100 mum HNE.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.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry, Vanderbilt University School of Medicine, Nashville, Tennessee 37232, USA.

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 mum 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 mum. 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.

Show MeSH
Related in: MedlinePlus