Protein oxidation and peroxidation

Abstract

Proteins are major targets for radicals and two-electron oxidants in biological systems due to their abundance and high rate constants for reaction. With highly reactive radicals damage occurs at multiple side-chain and backbone sites. Less reactive species show greater selectivity with regard to the residues targeted and their spatial location. Modification can result in increased side-chain hydrophilicity, side-chain and backbone fragmentation, aggregation via covalent cross-linking or hydrophobic interactions, protein unfolding and altered conformation, altered interactions with biological partners and modified turnover. In the presence of O2, high yields of peroxyl radicals and peroxides (protein peroxidation) are formed; the latter account for up to 70% of the initial oxidant flux. Protein peroxides can oxidize both proteins and other targets. One-electron reduction results in additional radicals and chain reactions with alcohols and carbonyls as major products; the latter are commonly used markers of protein damage. Direct oxidation of cysteine (and less commonly) methionine residues is a major reaction; this is typically faster than with H2O2, and results in altered protein activity and function. Unlike H2O2, which is rapidly removed by protective enzymes, protein peroxides are only slowly removed, and catabolism is a major fate. Although turnover of modified proteins by proteasomal and lysosomal enzymes, and other proteases (e.g. mitochondrial Lon), can be efficient, protein hydroperoxides inhibit these pathways and this may contribute to the accumulation of modified proteins in cells. Available evidence supports an association between protein oxidation and multiple human pathologies, but whether this link is causal remains to be established.

Keywords: UV; amino acid oxidation; hydroperoxides; peroxidation; peroxides; protein oxidation; radicals; singlet oxygen.

Figure 1. Examples of oxidant species (both two-electron oxidants and radicals) generated from activated leucocytes and their interconversion

Abbreviation: MPO, myeloperoxidase.

Figure 2. Abundance of potential targets for one- and two-electron oxidants in various biological systems, including plasma, liver and leucocytes

Replotted data from [7]: Davies, M.J. (2005) The oxidative environment and protein damage. Biochim. Biophys. Acta 1703, 93–109.

Figure 3. Overview of biological fates of carbon-centred (R^•, highlighted in red) and peroxyl radicals (ROO^•) in biological systems with subsequent formation of amino acid-, peptide- and protein-hydroperoxides (highlighted in blue)

Figure 4. Summary of currently known oxidation systems that can give rise to amino acid-, peptide- and protein-hydroperoxides in the presence of molecular oxygen (O₂)

This list is unlikely to be exhaustive.

Figure 5. Formation of hydroperoxides on reaction of Tyr phenoxyl radicals and Trp indolyl radicals with the superoxide radical anion, O₂^−•

In the case of the Tyr-derived species, these hydroperoxides can undergo further reactions with nucleophiles, including thiol, amine and amide groups to give more complex structures as a result of the presence of the conjugated double bond and carbonyl group, which is a reactive Michael acceptor. The resulting structures may retain the hydroperoxide function (see text).

Figure 6. Peroxidic species identified on reaction of singlet oxygen (¹O₂) with reactive methionine, cysteine, tyrosine, histidine and tryptophan side chains

Other species may also be formed, particularly with histidine (see text).

Figure 7. Overview of one- (radical) and two-electron (molecular) reactions of amino acid-, peptide- and protein-hydroperoxides (highlighted in blue)

The two-electron reactions occur predominantly with Cys residues to give sulfenic acids, disulfides, and higher oxy acids. Reaction has also been reported for Met, some disulfides such as lipoic acid (not shown) and selenium-containing compounds, including selenomethionine and selenocysteine (Sec)-containing enzymes such as thioredoxin reductase and glutathione peroxidase (not shown). One-electron reduction yields alkoxyl radicals and further oxidation reactions (for further details of specific mechanisms see Figure 8), whereas one-electron oxidation may yield peroxyl radicals; the latter process is poorly characterized.

Figure 8. Secondary fragmentation, rearrangement and hydrogen atom abstraction reactions of alkoxyl radicals (RO^•) generated from amino acid-, peptide- and protein-hydroperoxides

Decomposition of hydroperoxides (highlighted in blue) to RO^• can result in stable protein products (carbonyls and alcohols, highlighted in red), loss of protein side chains as low-molecular-mass carbonyls (highlighted in green), backbone fragmentation (highlighted in yellow), as well as further reactive radicals than can propagate damage and chain reactions.

Figure 9. Potential fates of hydroperoxides present on amino acids, peptides and proteins in biological systems

For further details see text.