Self-hydroxylation of taurine/α-ketoglutarate dioxygenase:: evidence for more than one oxygen activation mechanism
Self-hydroxylation of taurine/α-ketoglutarate dioxygenase:: evidence for more than one oxygen activation mechanism
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DOI：
10.1007/s00775-005-0059-4
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发表时间：
2006-01-01
影响因子：
3
通讯作者：
Que, L
中科院分区：
文献类型：
--
作者：
Koehntop, KD;Marimanikkuppam, S;Que, L
2-氨基乙磺酸（牛磺酸）/ α -酮戊二酸（α KG）双加氧酶（TauD）是一种单核非血红素铁酶，在O-2、α KG和Fe（II）存在下催化牛磺酸羟基化生成亚硫酸盐和氨基乙醛。Fe(II)TauD与aKG或琥珀酸盐（aKG的脱羧产物）络合，在没有主要底物的情况下与0反应，分别生成550 nm和720 nm的发色团，这些发色团可以通过添加或去除结合的碳酸氢盐而相互转换，并具有Fe(III)-儿茶酚酸络合物的共振拉曼特征。诱变研究表明，这两种反应都导致活性位点残基Tyr73的自羟基化，液相色谱纳米喷雾质谱/质谱证据证实了琥珀酸盐反应的这一结果。此外，同位素标记共振拉曼研究表明，加入酪氨酸残基的氧原子分别来自于(H2O)-O-18和O-18(2)的α - KG和琥珀酸盐反应，表明了不同的机制途径。鉴于α kg依赖的羟基化可能是通过已知在底物羟基化过程中产生的Fe(I-V) = O中间体进行的，我们提出Fe(III)-OOH（或Fe(V) = O）作为琥珀酸依赖反应中的氧化物质。这些结果证明了具有2- his -1-羧酸三聚体的酶的两种氧化机制，取决于电子源是否提供一个或两个电子。
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.