Radical Formation and Migration in Myoglobins

Abstract

Three EPR signals from individual free radical species have been identified in the EPR spectra of horse heart metmyoglobin (HH metMb) mixed with hydrogen peroxide (H2O2). The peroxyl radical EPR signal was assigned to the Trp14-OO•radical, the seven component signal – to the Tyr103•radical and the singlet EPR signal was assigned to the Tyr146•radical. Apo-Mb (haem free HH Mb) added in various concentrations to the native metMb prior to H2O2addition affected the yields of the three types of radicals. As the concentrations of metMb and H2O2were kept constant, the yield of the primary radical formed is the same in all experiments, H2O2being unable to produce any radical in the reaction with a haem free protein. Nevertheless, the addition of apo-Mb resulted in an increase of the Tyr146•radical concentration and in a quantitatively similar decrease of the Tyr103•radical concentration. These changes were dependent on the concentration of the added apo-Mb. Thus we show that a radical transfer Tyr103•→ Tyr146•occurs and that this reaction is protein concentration dependent. The question whether this radical transfer is inter- or intra-molecular is discussed. A similarity is drawn between the system studied and the sperm whale metMb/H2O2system, for which the radical transfer Tyr103•→ Tyr151•has been previously suggested.