Measurement of Heme Ruffling Changes in MhuD Using UV-vis Spectroscopy
Measurement of Heme Ruffling Changes in MhuD Using UV-vis Spectroscopy
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DOI：
10.1021/acs.jpcb.6b01497
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发表时间：
2016-04-28
影响因子：
3.3
通讯作者：
Liptak, Matthew D.
中科院分区：
文献类型：
--
作者：
Graves, Amanda B.;Graves, Max T.;Liptak, Matthew D.
几十年来，人们已经知道血红素的平面外褶皱畸变会干扰其紫外 - 可见吸收（Abs）光谱，但是增加的褶皱是导致索雷特带红移还是蓝移一直是一个有争议的话题。本文所介绍的结核分枝杆菌MhuD的光谱和计算特性解决了这一争议，MhuD是一种将血红素、氧气和还原当量转化为非血红素铁和藻胆素的酶。已使用氢 - 1核磁共振、吸收光谱和磁圆二色性光谱对W66F和W66A MhuD进行了表征，并且这些数据已被用于建立一个经过实验验证的褶皱态三价血红素理论模型。所建立的PBE密度泛函理论（DFT）模型准确地重现了从野生型酶观察到的光谱变化，并且通过分析电子结构的PBE DFT描述揭示了这些褶皱诱导变化的潜在量子力学起源。少量的血红素褶皱对Q带的能量没有影响，并且由于Fe 3d(xy)和卟啉a(2u)轨道的对称性允许混合而使索雷特带蓝移。较大程度的褶皱由于卟啉环内π键的破坏而使Q带和索雷特带均红移。
For decades it has been known that an out-of-plane ruffling distortion of heme perturbs its UV-vis absorption (Abs) spectrum, but whether increased ruffling induces a red or blue shift of the Soret band has remained a topic of debate. This debate has been resolved by the spectroscopic and computational characterization of Mycobacterium tuberculosis MhuD presented here, an enzyme that converts heme, oxygen, and reducing equivalents to nonheme iron and mycobilin. W66F and W66A MhuD have been characterized using H-1 nuclear magnetic resonance, Abs, and magnetic circular dichroism spectroscopies, and the data have been used to develop an experimentally validated theoretical model of ruffled, ferric heme. The PBE density functional theory (DFT) model that has been developed accurately reproduces the observed spectral changes from wild type enzyme, and the underlying quantum mechanical origins of these ruffling-induced changes were revealed by analyzing the PBE DFT description of the electronic structure. Small amounts of heme ruffling have no influence on the energy of the Q-band and blue-shift the Soret band due to symmetry-allowed mixing of the Fe 3d(xy) and porphyrin a(2u) orbitals. Larger amounts of ruffling red-shift both the Q and Soret bands due to disruption of pi-bonding within the porphyrin ring.