Self-hydroxylation of taurine/α-ketoglutarate dioxygenase:: evidence for more than one oxygen activation mechanism

2-Aminoethanesulfonic acid (taurine)/alpha-ketoglutarate (alpha KG) dioxygenase (TauD) is a mononuclear non-heme iron enzyme that catalyzes the hydroxylation of taurine to generate sulfite and aminoacetaldehyde in the presence Of O-2, alpha KG, and Fe(II). Fe(II)TauD complexed with aKG or succinate, the decarboxylated product of aKG, reacts with 0, in the absence of prime substrate to generate 550- and 720-nm chromophores, respectively, that are interconvertible by the addition or removal of bound bicarbonate and have resonance Raman features characteristic of an Fe(III)-catecholate complex. Mutagenesis studies suggest that both reactions result in the self-hydroxylation of the active-site residue Tyr73, and liquid chromatography nano-spray mass spectrometry/mass spectrometry evidence corroborates this result for the succinate reaction. Furthermore, isotope-labeling resonance Raman studies demonstrate that the oxygen atom incorporated into the tyrosyl residue derives from (H2O)-O-18 and O-18(2) for the alpha KG and succinate reactions, respectively, suggesting distinct mechanistic pathways. Whereas the alpha KG-dependent hydroxylation likely proceeds via an Fe(I-V) = O intermediate that is known to be generated during substrate hydroxylation, we propose Fe(III)-OOH (or Fe(V) = O) as the oxygenating species in the succinate-dependent reaction. These results demonstrate the two oxygenating mechanisms available to enzymes with a 2-His-1-carboxylate triad, depending on whether the electron source donates one or two electrons.