Tryptophanyl radical

The other amino acid residue present in proteins that is susceptible to oxidation is the indole moiety of tryptophan (Fig. 11). The reducing potential of tryptophan is considerably less than that of cysteine and methionine, so oxidation of tryptophanyl residues usually does not occur until all exposed thiol residues are oxidized. Also, the spontaneous oxidation of tryptophanyl residues in proteins is much less probable than that of cysteinyl and methionyl residues. Tryptophan residues are the only chromophoric moieties in proteins which can be photooxi-dized to tryptophanyl radicals by solar UV radiation, even by wavelengths as long as 305 nm (B12). Tryptophanyl residues readily react with all reactive oxygen species, hypochlorite, peroxynitrite, and chloramines. Oxidative modifications of other amino acid residues require use of strong oxidants, which eventually are produced in the cells. Detailed mechanisms of action of these oxidants is described in subsequent sections of this chapter. 

It is known that superoxide reacts very slowly with all amino-acids since all rate constants are below 100 mol l 1 s (89). Hence its reactivity with proteins without prosthetic group is low (89). One exception seems to be collagen, in which proline residues are oxidized into hydroxyproline (90). On the other hand, superoxide reacts efficiently with free radicals such as tryptophanyl radical (91). Reaction is fast with metalloproteins. It proceeds mostly by oxidizing or reducing the metal center. Some characteristics and rate constants of reactions with metalloproteins are given in table 7. It is obvious that products are often unknown and that the mechanism is sometimes unclear. It seems that there is no reaction with transferrin (92) and horseradish and lacto-peroxidase compounds II (93). The reason is unknown. 

In peptides and proteins, oxidation of tryptophan is followed by tryptophanyl radical reduction by tyrosine, leading to tyrosinyl radical. This reaction was shown first by Prutz and co-workers (120). Azide radicals are very convenient for this study. This process is easily visualized by pulse radiolysis since both free radicals absorb at different wavelengths (table 3) and the time scale for this reaction goes to microsecond for small peptides to millisecond for proteins. This reaction may occur by intramolecular step and thus it constitutes an excellent model to investigate long range intramolecular electron transfer. These results will be discussed further (see 5.1). 

Ivancich, A., P. Dorlet et al. (2001). Multifrequency high-field EPR study of the tryptophanyl and tyrosyl radical intermediates in wild-type and the W191G mutant of cytochrome c peroxidase. J. Am. Chem. Soc. 123 5050-5058. 

Recently, Batthyany et al. [133] pointed out that the reduction of cupric ions bound to apolipoprotein B-100 by endogenous LDL components might be an initiation step in copper-mediated LDL oxidation. They suggested that this reaction proceeds to form cuprous ion and the protein-tryptophanyl free radical the latter was identified on the basis of EPR spectrum with spin-trap 2-methyl-2-nitrosopropane. 

The realization of the widespread occurrence of amino acid radicals in enzyme catalysis is recent and has been documented in several reviews (52-61). Among the catalytically essential redox-active amino acids glycyl [e.g., anaerobic class III ribonucleotide reductase (62) and pyruvate formate lyase (63-65)], tryptophanyl [e.g., cytochrome peroxidase (66-68)], cysteinyl [class I and II ribonucleotide reductase (60)], tyrosyl [e.g., class I ribonucleotide reductase (69-71), photosystem II (72, 73), prostaglandin H synthase (74-78)], and modified tyrosyl [e.g., cytochrome c oxidase (79, 80), galactose oxidase (81), glyoxal oxidase (82)] are the most prevalent. The redox potentials of these protein residues are well within the realm of those achievable by biological oxidants. These redox enzymes have emerged as a distinct class of proteins of considerable interest and research activity. 

At physiological pH the protonated form of ONOO-, the peroxynitrous acid (ONOOH), is unstable and decomposes to nitrate (N03). ONOOH can also react directly with reductants or can decompose by homolytic dissociation to form nitrogen dioxide (N02) and hydroxyl radical (OH"), or can dissociate by a heterolytic mechanism to yield nitryl cation (NOj), which reacts with thiol, methionyl, tyro-syl, and tryptophanyl residues in proteins. 

Packer JE, Mahood JS, Willson RL et al (1981) Reactions of the trichloromethylperoxy free radical (CI3COO ) with tryptophan, tryptophanyl-tyiosine and lysozyme. Int J Radiat Biol 39 135-141... 

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