Tyrosyl radical generated by myeloperoxidase catalyzes the oxidative cross-linking of proteins.

Abstract

Phagocytes generate H202 for use by a secreted heme enzyme, myeloperoxidase, to kill invading bacteria, viruses, and fungi. We have explored the possibility that myeloperoxidase might also convert L-tyrosine to a radical catalyst that cross-links proteins. Protein-bound tyrosyl residues exposed to myeloperoxidase, H202, and L-tyrosine were oxidized to oo'-dityrosine, a stable product of the tyrosyl radical. The cross-linking reaction required L-tyrosine but was independent of halide and free transition metal ions; the heme poisons azide and aminotriazole were inhibitory. Activated neutrophils likewise converted polypeptide tyrosines to dityrosine. The pathway for oxidation of peptide tyrosyl residues was dependent upon L-tyrosine and was inhibited by heme poisons and catalase. Dityrosine synthesis was little affected by plasma concentrations of Cl-and amino acids, suggesting that the reaction pathway might be physiologically relevant. The requirement for free L-tyrosine and H202 for dityrosine formation and the inhibition by heme poisons support the hypothesis that myeloperoxidase catalyzes the cross-linking of proteins by a peroxidative mechanism involving tyrosyl radical. In striking contrast to the pathways generally used to study protein oxidation in vitro, the reaction does not require free metal ions. We speculate that protein dityrosine cross-linking by myeloperoxidase may play a role in bacterial killing or injuring normal tissue. The intense fluorescence and stability of biphenolic compounds may allow dityrosine to act as a marker for proteins oxidatively damaged by myeloperoxidase in phagocyte-rich inflammatory lesions. (J.Clin. Invest. 1993Invest. . 91:2866Invest. -2872

Detection of HOCl-mediated protein oxidation products in the extracellular matrix of human atherosclerotic plaques

“…Previous studies have implicated the generation of Tyr phenoxyl radicals generated by MPO in the formation of di-Tyr [26,53]. The elevated levels of di-Tyr observed in plaque ECM are therefore consistent with the close juxtaposition of this enzyme with the ECM of arterial lesions.…”

Production of Brominating Intermediates by Myeloperoxidase

“…Myeloperoxidase and Eosinophil Peroxidase-Myeloperoxidase (A 430 / A 280 Ͼ 0.8) was isolated from HL-60 cells by sequential lectin affinity, ion exchange, and size exclusion chromatographies (43,44). Enzyme concentration was determined spectrophotometrically (⑀ 430 ϭ 178 mM Peroxidase Activity Assay-The purity of myeloperoxidase and eosinophil peroxidase were assessed by peroxidase activity using nondenaturing polyacrylamide slab gel electrophoresis and gel system 8 (46,47).…”

Biochemistry and pathology of radical-mediated protein oxidation

“…During metal-ion and radiolytic attack, lipid and protein oxidation are often concurrent, and occur even while much vitamin E remains in the particles. In contrast, hypochlorite selectively attacks the protein, consuming mainly lysine, tryptophan, cysteine and methionine residues, and giving rise to chloramines [97,150]. Exogenous tyrosine can be cross-linked to apoB tyrosines by hypochlorite [150], although it is not clear whether this occurs with physiological levels of tyrosine.…”