Nitrosation of N-terminally blocked tryptophan and tryptophan-containing peptides by peroxynitrite.
Tryptophan is known to be a major target of oxidative stress and to take part in electron transfer. In proteins, its fluorescence is extinguished after treatment with oxidative agents, like peroxynitrite (ONOO(-)/ONOOH) - the product of the reaction of NO* and superoxide anion (O*(2)(-)) radicals. The main reactions of N-blocked tryptophan derivatives (melatonin or N-acetyl-L-tryptophan) exposed to peroxynitrite at physiological pH are oxidation to formylkynuramine or formylkynurenine, respectively, and nitrosation, which leads to substituted 1-nitrosoindoles. Here we show that peroxynitrite-induced nitrosation is specific to N-blocked L-tryptophan derivatives and is not obtained with free L-tryptophan. Such a nitrosation can be evaluated by using 4,5-diaminofluorescein (DAF-2), which is converted to the fluorescent triazolofluorescein by NO* donors and nitrosating agents. N-acetyl-L-tryptophan was shown to be twice as efficient as melatonin in transferring NO from peroxynitrite to DAF-2. DAF-2 responses were then used to assess the ability of a series of L-tryptophan-containing peptides to give transient N-nitrosoindoles upon treatment with peroxynitrite. Many peptides proved not to be susceptible to nitrosation under these conditions. However, the N-terminally blocked peptide of endothelin-1 (Ac-Asp-Ile-Ile-Trp) reacted in a very similar fashion to melatonin; this shows that tryptophan residue nitrosation could occur when it was exposed to peroxynitrite.
Peyrot
Fabienne
F
Institut de Chimie des Substances Naturelles, CNRS, Avenue de la Terrasse, 91198 Gif-sur-Yvette, France.
Ducrocq
Claire
C
D019793:Fluorescein / Q000737:chemistry; D007211:Indoles / Q000737:chemistry; D008969:Molecular Sequence Data; D015394:Molecular Structure; D015538:Nitrosation; D010455:Peptides / Q000737:chemistry*; D030421:Peroxynitrous Acid / Q000737:chemistry*; D014364:Tryptophan / Q000737:chemistry*
C111205:4,5-diaminofluorescein; D007211:Indoles; D010455:Peptides; D030421:Peroxynitrous Acid; D014364:Tryptophan; D019793:Fluorescein