Most of the assays for detection of carbonylated proteins, the most general and widely used marker of severe protein oxidation, involve derivatization of the carbonyl group with 2,4-dinitrophenylhydrazine (DNPH), which leads to formation of a stable dinitrophenyl hydrazone product. Here, by using a Cys-containing model peptide and high-resolution mass spectrometry, we demonstrate that DNPH is not exclusively selective for carbonyl groups, because it also reacts with sulfenic acids, forming a DNPH adduct, through the acid-catalyzed formation of a thioaldehyde intermediate that is further converted to an aldehyde. β-Mercaptoethanol prevents the formation of the DNPH derivative because it reacts with the oxidized Cys residue, forming the corresponding disulfide.
|Titolo:||Protein carbonylation: 2,4-dinitrophenylhydrazine reacts with both aldehydes/ketones and sulfenic acids|
|Parole Chiave:||2,4-Dinitrophenylhydrazine; Cysteine sulfenic acid; Cysteine sulfinic acid; Cysteine sulfonic acid; Dinitrophenyl hydrazone adducts; Free radicals; Hydrogen peroxide; Mass spectrometry; Protein carbonyls; Thioaldehyde|
|Settore Scientifico Disciplinare:||Settore CHIM/08 - Chimica Farmaceutica|
Settore BIO/06 - Anatomia Comparata e Citologia
|Data di pubblicazione:||2009|
|Digital Object Identifier (DOI):||10.1016/j.freeradbiomed.2009.02.024|
|Appare nelle tipologie:||01 - Articolo su periodico|