Radiolysis Studies of Oxidation and Nitration of Tyrosine and Some Other Biological Targets by Peroxynitrite-Derived Radicals

Abstract

The widespread interest in free radicals in biology extends far beyond the effects of ionizing radiation, with recent attention largely focusing on reactions of free radicals derived from peroxynitrite (i.e., hydroxyl, nitrogen dioxide, and carbonate radicals). These radicals can easily be generated individually by reactions of radiolytically-produced radicals in aqueous solutions and their reactions can be monitored either in real time or by analysis of products. This review first describes the general principles of selective radical generation by radiolysis, the yields of individual species, the advantages and limitations of either pulsed or continuous radiolysis, and the quantitation of oxidizing power of radicals by electrode potentials. Some key reactions of peroxynitrite-derived radicals with potential biological targets are then discussed, including the characterization of reactions of tyrosine with a model alkoxyl radical, reactions of tyrosyl radicals with nitric oxide, and routes to nitrotyrosine formation. This is followed by a brief outline of studies involving the reactions of peroxynitrite-derived radicals with lipoic acid/dihydrolipoic acid, hydrogen sulphide, and the metal chelator desferrioxamine. For biological diagnostic probes such as 'spin traps' to be used with confidence, their reactivities with radical species have to be characterized, and the application of radiolysis methods in this context is also illustrated.

Keywords: DMPO; carbonate radicals; desferrioxiamine; dityrosine; hydrogen sulfide; hydroxyl radicals; lipoic acid; nitric oxide; nitrogen dioxide radicals; nitrotyrosine; radiolysis; tyrosine.

Figure 1

Pathways by which superoxide and nitric oxide can be converted to other oxidants in biology. The formation of peroxynitrite is emphasized. MPO, myeloperoxidase; AscH⁻, ascorbate.

Figure 2

Upper: examples of transient absorption spectra of CO₃^•– generated following pulse radiolysis of a solution of 40 µM N-t-BOC-L-tyrosine tert-butyl ester (BTBE) incorporated with liposomes prepared with egg yolk phosphatidyl choline at pH 8, 100 µs (filled circles) and 500 µs (empty circles) after a 15 Gy, 500 ns pulse (note the formation of absorption at 405 nm indicative of the formation of TyrO^•). Lower: example transients showing decay of the absorption (A) of CO₃^•–; (a) with no BTBE and (b) with 40 µM BTBE present.

Figure 3

Pathways of free-radical reactions with tyrosine which can be monitored by pulse radiolysis or HPLC (with absorbance, fluorescence and mass spectrometric detection) following steady-state radiolysis. Chemical structures of products showed in the figure can be found in the following references [7,56,57,58,59,60,61]. A putative structure of a tyrosine-glutathione adducts formed from the reaction of the TyrO^•-GS^• radical intermediates (dashed arrow) has been inferred from [53].