| Autor: |
Ehrenshaft M; Laboratory of Pharmacology, National Institute of Environmental Health Sciences, National Institutes of Health, Research Triangle Park, NC 27709, USA. ehrensh1@niehs.nih.gov, Silva SO, Perdivara I, Bilski P, Sik RH, Chignell CF, Tomer KB, Mason RP |
| Jazyk: |
angličtina |
| Zdroj: |
Free radical biology & medicine [Free Radic Biol Med] 2009 May 01; Vol. 46 (9), pp. 1260-6. |
| DOI: |
10.1016/j.freeradbiomed.2009.01.020 |
| Abstrakt: |
Reactions of tryptophan residues in proteins with radical and other oxidative species frequently lead to cleavage of the indole ring, modifying tryptophan residues into N-formylkynurenine (NFK) and kynurenine. Tryptophan modification has been detected in physiologically important proteins and has been associated with a number of human disease conditions. Modified residues have been identified through various combinations of proteomic analyses, tryptic digestion, HPLC, and mass spectrometry. Here we present a novel, immunological approach using polyclonal antiserum for detection of NFK. The specificity of our antiserum is confirmed using photooxidation and radical-mediated oxidation of proteins with and without tryptophan residues. The sensitivity of our antiserum is validated through detection of NFK in photooxidized myoglobin (two tryptophan residues) and in carbonate radical-oxidized human SOD1, which contains a single tryptophan residue. Analysis of photooxidized milk also shows that our antiserum can detect NFK residues in a mixture of proteins. Results from mass spectrometric analysis of photooxidized myoglobin samples corroborate the immunological data, detecting an increase in NFK content as the extent of photooxidation increases. |
| Databáze: |
MEDLINE |
| Externí odkaz: |
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