tailieunhanh - Báo cáo khoa học: Peroxidase catalyzed nitration of tryptophan derivatives Mechanism, products and comparison with chemical nitrating agents

The enzymatic nitration of tryptophan derivatives by oxi-dation of nitrite has been studied using lactoperoxidase and horseradish peroxidase, and compared with the chemical nitration produced by nitrogen dioxide and peroxynitrite. HPLC, mass spectra and NMR analysis of the mixture of products clearly show that nitrationoccurs at position 4-, 6-, 7-, andN 1 of the indole ring, and nitrosation at positionN 1 . Kinetic studies performed on peroxidase/NO2 – /H2O2 sys-tems showed substrate saturation behavior with all the tryptophan derivatives employed. . | Eur. J. Biochem. 271 2841-2852 2004 FEBS 2004 doi Peroxidase catalyzed nitration of tryptophan derivatives Mechanism products and comparison with chemical nitrating agents Alberto Sala Stefania Nicolis Raffaella Roncone Luigi Casella and Enrico Monzani Dipartimento di Chimica Generale Universita di Pavia Italy The enzymatic nitration of tryptophan derivatives by oxidation of nitrite has been studied using lactoperoxidase and horseradish peroxidase and compared with the chemical nitration produced by nitrogen dioxide and peroxynitrite. HPLC mass spectra and NMR analysis of the mixture of products clearly show that nitration occurs at position 4- 6- 7- and N 1 of the indole ring and nitrosation at position N1. Kinetic studies performed on peroxidase NO- H2O2 systems showed substrate saturation behavior with all the tryptophan derivatives employed. The rate dependence on nitrite concentration was found to be linear with horseradish peroxidase while it exhibited saturation behavior with lactoperoxidase. The composition of the product mixture depends on the nitrating agent. While the production of 4-nitro 6-nitro 7-nitro and N1-nitro derivatives follows a similar trend indicating that they are formed according to a similar mechanism the ratio between the N1-nitroso derivative and other derivatives depends markedly on the nitrite concentration when tryptophan modification is performed by the peroxidase H2O2 nitrite systems. Analysis of the data indicates that at low nitrite concentration the enzymatic reaction occurs through the classical peroxidase cycle. At high nitrite concentration the reaction proceeds through a different intermediate that we assume to be a protein bound peroxynitrite species. Keywords heme proteins nitration peroxidases peroxynitrite tryptophan. Reactive nitrogen species RNS are formed continuously in vivo but their concentration in the body rises notably during inflammation and several pathophysiological conditions