HEME PROTEIN RADICALS - FORMATION, FATE, AND BIOLOGICAL CONSEQUENCES

The oxidation of myoglobin by H2O2 yields ferrylmyoglobin, which conta ins two oxidizing equivalents: the oxoferryl complex and an amino acid radical. This study examines the electron paramagnetic resonance (EPR ) properties of the resulting amino acid radicals and their inherent k inetic features at [H2O2]/[protein] ratios close to physiological cond itions (i.e., less than or equal to 1). The EPR spectrum obtained with continuous flow at room temperature consisted of a composite of three signals: a low intensity signal and two high intensity signals. The f ormer had a g-value of 2.014, contributed 10-15% to the overall spectr um and was ascribed to a peroxyl radical. Of the two high intensity si gnals, one consisted of a six-line spectrum (g = 2.0048) that contribu ted approximately 17-19% to the overall signal; hyperfine splitting co nstants to ring protons permitted to identify this signal as a tyrosyl radical. The other high intensity signal (with similar g-value and un derlying that of the tyrosyl radical) was ascribed to an aromatic amin o acid upon comparison with the EPR characteristics for radicals in ar omatic amino acid-containing peptides. Analysis of these data in conne ction with amino acid analysis and the EPR spectra obtained under simi lar conditions with another hemoprotein, hemoglobin, allowed to sugges t a mechanism for the formation of the protein radicals in myoglobin. The aromatic amino acid radical was observed to be relatively long liv ed in close proximity to the heme iron. Hence, it is likely that this is the first site of protein radical; reduction of the oxoferryl compl ex by Tyr (Fe-IV=O + Tyr-OH + H+ --> Fe-III + H2O + Tyr-O-.)-and alter natively by other amino acids-leads to the subsequent formation of oth er amino acid radicals within an electron-transfer process throughout the protein. This view suggests that the protein radical(s) is highly delocalized within the globin moiety in a dynamic process encompassing electron tunneling through the backbone chain or H-bonds;md leading t o the formation of secondary radicals. (C) 1998 Elsevier Science Inc.