Histidine Orientation Modulates the Structure and Dynamics of a de Novo Metalloenzyme Active Site.
Histidine Orientation Modulates the Structure and Dynamics of a de Novo Metalloenzyme Active Site.
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组氨酸取向调节从头脂酶活性位点的结构和动力学。
DOI：
10.1021/jacs.5b02840
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发表时间：
2015-08-19
影响因子：
15
通讯作者：
Kubarych KJ
中科院分区：
文献类型：
--
作者：
Ross MR;White AM;Yu F;King JT;Pecoraro VL;Kubarych KJ
利用二维红外光谱对金属酶活性位点的超快动力学进行了监测。平行的螺旋状α-螺旋的同型三聚体含有一个His3-Cu(I)金属位点，CO在此结合，并作为自组装配合物疏水内部的振动探针。Cu-CO的超快光谱动力学揭示了CO的振动激发引发的前所未有的超快（2ps）非平衡结构重排。这一初始快速阶段之后是天然蛋白质中典型的金属-CO振动的慢得多的~ 40ps的振动弛豫。为了确定隐藏的偶联坐标，分别通过QM和QM/MM计算对小分子类似物和全肽进行了研究。计算表明，组氨酸在配位Cu时的二面角变化控制着CO的拉伸与Cu - c - o弯曲坐标之间的耦合。对不同优化结构在CO配体位置具有显著不同的静电场强度的分析表明，拉伸-弯曲耦合的起源不是直接由穿过空间的静电引起的。相反，组氨酸侧链的~3.6 D大偶极矩有效地将静电环境转换为局部金属配位取向。第一配位球对蛋白质静电的敏感性及其在改变结合配体表面势能中的作用表明，远程静电可以通过酶的设计来微调功能。
The ultrafast dynamics of a de novo metalloenzyme active site is monitored using two-dimensional infrared spectroscopy. The homotrimer of parallel, coiled coil α-helices contains a His3-Cu(I) metal site where CO is bound and serves as a vibrational probe of the hydrophobic interior of the self-assembled complex. The ultrafast spectral dynamics of Cu-CO reveals unprecedented ultrafast (2 ps) nonequilibrium structural rearrangements launched by vibrational excitation of CO. This initial rapid phase is followed by much slower ~40 ps vibrational relaxation typical of metal-CO vibrations in natural proteins. To identify the hidden coupled coordinate, small molecule analogues and the full peptide were studied by QM and QM/MM calculations, respectively. The calculations show that variation of the histidines’ dihedral angles in coordinating Cu controls the coupling between the CO stretch and the Cu–C–O bending coordinates. Analysis of different optimized structures with significantly different electrostatic field magnitudes at the CO ligand site indicates that the origin of the stretch–bend coupling is not directly due to through-space electrostatics. Instead, the large, ~3.6 D dipole moments of the histidine side chains effectively transduce the electrostatic environment to the local metal coordination orientation. The sensitivity of the first coordination sphere to the protein electrostatics and its role in altering the potential energy surface of the bound ligands suggests that long-range electrostatics can be leveraged to fine-tune function through enzyme design.