| Abstract
| - Taurine/α-ketoglutarate dioxygenase (TauD), a member of the broad class of non-hemeFe(II) oxygenases, converts taurine (2-aminoethanesulfonate) to sulfite and aminoacetaldehyde whiledecomposing α-ketoglutarate (αKG) to form succinate and CO2. Under anaerobic conditions, the additionof αKG to Fe(II)TauD results in the formation of a broad absorption centered at 530 nm. On the basis ofstudies of other members of the αKG-dependent dioxygenase superfamily, we attribute this spectrum tometal chelation by the substrate C-1 carboxylate and C-2 carbonyl groups. Subsequent addition of taurineperturbs the spectrum to yield a 28% greater intensity, an absorption maximum at 520 nm, and distinctshoulders at 480 and 570 nm. This spectral change is specific to taurine and does not occur when2-aminoethylphosphonate or N-phenyltaurine is added. Titration studies demonstrate that each TauD subunitbinds a single molecule of Fe(II), αKG, and taurine. In addition, these studies indicate that the affinity ofTauD for αKG is enhanced by the presence of taurine. α-Ketoadipate, the other α-keto acid previouslyshown to support TauD activity, and α-ketocaproate lead to the formation of weak 520 nm-like spectrawith Fe(II)TauD in the presence of taurine; however, corresponding spectra at 530 nm are not observedin the absence of taurine. Pyruvate and α-ketoisovalerate fail to elicit absorption bands in this region ofthe spectrum, even in the presence of taurine. Stopped-flow UV−visible spectroscopy reveals that the530 and 520 nm spectra associated with αKG−Fe(II)TauD and taurine−αKG−Fe(II)TauD are formed atcatalytically competent rates (∼40 s-1). The rate of chromophore formation was independent of substrateor enzyme concentration, suggesting that αKG binds to Fe(II)TauD prior to the formation of a chromophoricspecies. Significantly, the taurine−αKG−Fe(II)TauD state, but not the αKG−Fe(II)TauD species, reactsrapidly with oxygen (42 ± 9 s-1). Using the data described herein, we develop a preliminary kineticmodel for TauD catalysis.
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