Peroxynitrite is a reactive nitrogen species linked to numerous inflammation-mediated disease states. Precursors, superoxide and nitric oxide, are generated from independent sources and react to generate peroxynitrite at a near diffusion-controlled rate. Unfortunately, just as quickly as it forms this reactive species decomposes. For this reason, synthesis of peroxynitrite in situ has proven to be complicated and tedious. To address this obstacle we have engineered a two-component light-controlled peroxynitrite-generating platform that evolves sustained levels of peroxynitrite under physiological conditions.   In order to establish the utility of our system as a bioanalytical tool we investigated the hallmark reaction of peroxynitrite with tyrosine to from 3-nitrotyrosine. This post-translational modification has been proposed as a footprint of peroxynitrite in vivo; however, controversy has surrounded this reaction due to varied results in the presence of co-generated and chemical peroxynitrite sources. In the wells of our platform we studied the peroxynitrite-mediated nitration of free tyrosine and tyrosine-containing model peptides under varied pH and CO2 concentrations and in the presence of thiols. Interestingly, we found that 3-nitrotyrosine is in fact the predominant species formed when tyrosine is exposed to peroxynitrite. Further, the extent of nitration observed within the wells of our platform are significantly greater than those observed in the presence of chemical sources of peroxynitrite. The results of this study highlight the potential of this platform to uncover the complex chemistry of peroxynitrite.

Engineering a two-component light-controlled peroxynitrite generating platform: A new tool to study the chemistry of peroxynitrite

248th National Meeting (2014)

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