Journal article
Modification of heat shock protein 90 by 4-hydroxynonenal in a rat model of chronic alcoholic liver disease
The Journal of pharmacology and experimental therapeutics, Vol.315(1), pp.8-15
10/2005
DOI: 10.1124/jpet.105.088088
PMID: 15951401
Abstract
Lipid peroxidation during oxidative stress leads to increased concentrations of thiol-reactive alpha,beta-unsaturated aldehyde, including 4-hydroxy-2-nonenal (4-HNE) and 4-oxo-2-nonenal (4-ONE). These aldehydes have a documented ability to disrupt protein function following adduct formation with specific residues. Therefore, to identify 4-HNE-modified proteins in a model of ethanol-induced oxidative stress, a proteomic approach was applied to liver fractions prepared from rats fed a combination high-fat/ethanol diet. The results revealed that essential 90-kDa heat shock protein (Hsp90) was consistently modified by 4-HNE in the alcohol-treated animals. In vitro chaperoning experiments using firefly luciferase as a client protein were then performed to assess the functional effect of 4-HNE modification on purified recombinant human Hsp90, modified with concentrations of this aldehyde ranging from 23 to 450 microM. Modification of Hsp90 with 4-ONE also led to significant inhibition of the chaperone. Because 4-HNE and 4-ONE react selectively with Cys, a thiol-specific mechanism of inhibition was suggested by these data. Therefore, thiol sensitivity was confirmed following treatment of Hsp90 with the specific thiol modifier N-ethylmaleimide, which resulted in more than 99% inactivation of the chaperone by concentrations as low as 6 microM (1:1 M ratio). Finally, tryptic digest of 4-HNE-modified Hsp90 followed by liquid chromatography/tandem mass spectrometry peptide analysis identified Cys 572 as a site for 4-HNE modification. The results presented here thus establish that 4-HNE consistently modifies Hsp90 in a rat model of alcohol-induced oxidative stress and that the chaperoning activity of this protein is subject to dysregulation through thiol modification.
Details
- Title: Subtitle
- Modification of heat shock protein 90 by 4-hydroxynonenal in a rat model of chronic alcoholic liver disease
- Creators
- David L Carbone - Department of Pharmaceutical Sciences, School of Pharmacy, University of Colorado Health Sciences Center, Denver, CO 80262, USAJonathan A DoornZachary KieblerBrian R IckesDennis R Petersen
- Resource Type
- Journal article
- Publication Details
- The Journal of pharmacology and experimental therapeutics, Vol.315(1), pp.8-15
- Publisher
- United States
- DOI
- 10.1124/jpet.105.088088
- PMID
- 15951401
- ISSN
- 0022-3565
- eISSN
- 1521-0103
- Grant note
- R01AA09300 / NIAAA NIH HHS F31 AA014308 / NIAAA NIH HHS F32 ES11937 / NIEHS NIH HHS
- Language
- English
- Date published
- 10/2005
- Academic Unit
- Iowa Neuroscience Institute; Pharmaceutical Sciences and Experimental Therapeutics; Medicinal and Natural Products Chemistry
- Record Identifier
- 9984070141402771
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