Efficient identification and quantification of peptides containing nitrotyrosine by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry after derivatization.
Protein nitration at tyrosine residues proceeds to form 3-nitrotyrosine; this product is today used as a biomarker of nitrative stress. We have developed an efficient method with which to identify nitrated peptides and to quantify protein nitration levels in different biological samples by a combination of the chemical derivatization of nitrotyrosine-containing peptides and mass spectrometry. Our strategy includes: 1) the protection of both N-terminal amines and epsilon-amines of lysine residues by acetylation with (13)C(0)/(13)C(4)- or D(0)/D(6)-acetic anhydride; 2) the reduction of nitrotyrosine to aminotyrosine with sodium hydrosulfite; and 3) the derivatization of aminotyrosine with 1-(6-methyl[D(0)/D(3)]nicotinoyloxy)succinimide. The utility of our method is demonstrated with nitrotyrosine-containing angiotensin II and bovine serum albumin as the model compounds.
Tsumoto
Hiroki
H
Research Institute of Pharmaceutical Sciences, Musashino University, Tokyo, Japan.
Taguchi
Ryo
R
Kohda
Kohfuku
K
D000595:Amino Acid Sequence; D000804:Angiotensin II / Q000737:chemistry*; D000818:Animals; D002417:Cattle; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry*; D012710:Serum Albumin, Bovine / Q000737:chemistry*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods*; D014443:Tyrosine / Q000031:analogs & derivatives* / Q000032:analysis
D010455:Peptides; D000804:Angiotensin II; D012710:Serum Albumin, Bovine; C002744:3-nitrotyrosine; D014443:Tyrosine