Resonance Raman study on the oxygenated and the ferryl-oxo species of indoleamine 2,3-dioxygenase during catalytic turnover

Resonance Raman (RR) spectra of the oxygenated and Fe-IV=O reaction intermediates of indoleamine 2,3-dioxygenase (IDO) are reported. Absorption and RR spectra reveal that the electronic and geometric structures of the two respective species at pH 6.5 and pH 8.0 are the same, although the enzymatic activity at pH 6.5 is 6 times higher than at pH 8.0. The results thus further support our current understanding that the Fe-IV=O heme species is the active species in the IDO reaction cycle, although its presence was unexpected. The Fe-O-2 and the O-O stretching frequencies of the IDO-Trp-O-2 ternary complex at Trp concentrations of 50 mu M and 8 mM are essentially identical. These results suggest that "substrate inhibition'' of enzymatic activity occurs by binding of a second substrate molecule to an unknown binding site and not to the heme pocket.