title: Protein carbonylation: 2,4-dinitrophenylhydrazine reacts with both aldehydes/ketones and sulfenic acids.
abstract: 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. beta-Mercaptoethanol prevents the formation of the DNPH derivative because it reacts with the oxidized Cys residue, forming the corresponding disulfide.
authors.affiliation: Department of Biology, University of Milan, Italy. quack@unimi.it
authors.forename: Isabella
authors.initials: I
authors.lastname: Dalle-Donne
authors.forename: Marina
authors.initials: M
authors.lastname: Carini
authors.forename: Marica
authors.initials: M
authors.lastname: Orioli
authors.forename: Giulio
authors.initials: G
authors.lastname: Vistoli
authors.forename: Luca
authors.initials: L
authors.lastname: Regazzoni
authors.forename: Graziano
authors.initials: G
authors.lastname: Colombo
authors.forename: Ranieri
authors.initials: R
authors.lastname: Rossi
authors.forename: Aldo
authors.initials: A
authors.lastname: Milzani
authors.forename: Giancarlo
authors.initials: G
authors.lastname: Aldini
mesh_terms: D000447:Aldehydes / Q000737:chemistry*; D019295:Computational Biology; D003545:Cysteine / Q000737:chemistry / Q000378:metabolism; D019281:Dimerization; D007659:Ketones / Q000737:chemistry*; D008962:Models, Theoretical; D010084:Oxidation-Reduction; D010455:Peptides / Q000737:chemistry*; D010659:Phenylhydrazines / Q000737:chemistry* / Q000378:metabolism; D011485:Protein Binding; D050050:Protein Carbonylation; D013434:Sulfenic Acids / Q000737:chemistry*
chemical_list: D000447:Aldehydes; D007659:Ketones; D010455:Peptides; D010659:Phenylhydrazines; D013434:Sulfenic Acids; C004787:2,4-dinitrophenylhydrazine; D003545:Cysteine