ASSIGNMENT OF THE SPECTRA OF PROTEIN RADICALS IN CYTOCHROME-C PEROXIDASE

In oxidized cytochrome c peroxidase a peak at 570 nm has been attributed to an intermediate tryptophan free radical of indeterminate protonation state. Ab initio calculations of the spectra of the neutral and cationic free radicals of the indole side chain of tryptophan are used to assign the absorption spectra to the neutral radical. Earlier assignment attempts were confused by the large blue shift in water of the in vacuo transition. The relevant calculated transitions of both the neutral and cationic indole radicals, in fact, shift substantially to the blue in aqueous solution. The theoretical cation spectra do not agree with the experimental spectra and suggest a tautomeric form is observed. Calculations of the valence singlet and triplet excited states of benzene and triplet states of indole are also presented to support the ab initio calculations of the indole radical. Spin densities are calculated for both neutral and cation radicals. The small spin density on nitrogen found for the cation is more in agreement with experimental ENDOR data. The visible and ENDOR data suggest different states of protonation and cannot be reconciled to the electronic properties of the ground-state radical tautomers.