Probing the iron-substrate orientation for taurine/α-ketoglutarate dioxygenase using deuterium electron spin echo envelope modulation spectroscopy
Probing the iron-substrate orientation for taurine/α-ketoglutarate dioxygenase using deuterium electron spin echo envelope modulation spectroscopy
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DOI：
10.1021/bi700562t
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发表时间：
2007-05-22
期刊：
影响因子：
2.9
通讯作者：
McCracken, John
中科院分区：
文献类型：
--
作者：
Muthukumaran, Rajendra Bose;Grzyska, Piotr K.;McCracken, John
用电子自旋回波包络调制(ESEEM)谱研究了底物牛磺酸与牛磺酸/α-酮戊二酸(α-KG)双加氧酶(TauD)的非血红素Fe(II)中心之间的结构关系。研究是在TauD样品上进行的，用NO、共底物α-KG和质子化或特定的氚化牛磺酸处理。模拟回波ESEEM数据被分割，以消除来自H-1和N-14调制的干扰，并增强来自H-2的调制。对于在C-1位(与磺酸基相邻)的牛磺酸，H-2 ESEEM谱显示了来自单个氘核的偶极-偶极和氚核四极相互作用的特征。对牛磺酸在C-1和C-2上的氢化进行的平行测量表明，在重氢Larmor频率处有一个额外的ESEEM特征。在g=4到g=2范围内对这些数据的分析使我们能够确定底物牛磺酸相对于Fe(II)-NO，S=(3)/(2)顺磁中心磁轴的取向。根据以前的X射线结晶学研究和这一家族酶的催化机理，对这些结果进行了讨论。
The structural relationship between substrate taurine and the non-heme Fe(II) center of taurine/alpha-ketoglutarate (alpha KG) dioxygenase (TauD) was measured using electron spin echo envelope modulation (ESEEM) spectroscopy. Studies were conducted on TauD samples treated with NO, cosubstrate alpha KG, and either protonated or specifically deuterated taurine. Stimulated echo ESEEM data were divided to eliminate interference from H-1 and N-14 modulations and accentuate modulations from H-2. For taurine that was deuterated at the C-1 position (adjacent to the sulfonate group), H-2 ESEEM spectra show features that arise from dipole-dipole and deuterium nuclear quadrupole interactions from a single deuteron. Parallel measurements taken for taurine deuterated at both C-1 and C-2 show an additional ESEEM feature at the deuterium Larmor frequency. Analysis of these data at field positions ranging from g = 4 to g = 2 have allowed us to define the orientation of substrate taurine with respect to the magnetic axes of the Fe(II)-NO, S = (3)/(2), paramagnetic center. These results are discussed in terms of previous X-ray crystallographic studies and the proposed catalytic mechanism for this family of enzymes.