Department of Life Sciences, University of Modena and Reggio Emilia, via Campi 103, 41125 Modena, Italy.
Department of Chemistry and Geology, University of Modena and Reggio Emilia, via Campi 103, 41125 Modena, Italy.
J Inorg Biochem. 2018 Jan;178:70-86. doi: 10.1016/j.jinorgbio.2017.10.005. Epub 2017 Oct 16.
Neuroglobin is a monomeric globin containing a six-coordinate heme b, expressed in the nervous system, which exerts an important neuroprotective role. In the human protein (hNgb), Cys46 and Cys55 form an intramolecular disulfide bond under oxidizing conditions, whose cleavage induces a helix-to-strand rearrangement of the CD loop that strengthens the bond between the heme iron and the distal histidine. Hence, it is conceivable that the intramolecular disulfide bridge modulates the functionality of human neuroglobin by controlling exogenous ligand binding. In this work, we investigated the influence of the Cys46/Cys55 disulfide bond on the redox properties and on the pH-dependent conformational equilibria of hNgb, using UV-vis spectroelectrochemistry, cyclic voltammetry, electronic absorption spectroscopy and magnetic circular dichroism (MCD). We found that the SS bridge significantly affects the heme Fe(III) to Fe(II) reduction enthalpy (ΔH°') and entropy (ΔS°'), mostly as a consequence of changes in the reduction-induced solvent reorganization effects, without affecting the axial ligand-binding interactions and the polarity and electrostatics of the heme environment. Between pH3 and 12, the electronic properties of the heme of ferric hNgb are sensitive to five acid-base equilibria, which are scarcely affected by the Cys46/Cys55 disulfide bridge. The equilibria occurring at extreme pH values induce heme release, while those occurring between pH5 and 10 alter the electronic properties of the heme without modifying its axial coordination and low spin state. They involve the sidechains of non-coordinating aminoacids close to the heme and at least one heme propionate.
神经球蛋白是一种单体球蛋白，含有一个六配位的血红素 b，在神经系统中表达，发挥着重要的神经保护作用。在人类蛋白（hNgb）中，Cys46 和 Cys55 在氧化条件下形成一个分子内二硫键，其断裂诱导 CD 环的螺旋到链的重排，从而增强血红素铁与远端组氨酸之间的键。因此，可以想象分子内二硫键通过控制外源配体结合来调节人类神经球蛋白的功能。在这项工作中，我们使用紫外可见光谱电化学、循环伏安法、电子吸收光谱和圆二色性（MCD）研究了 Cys46/Cys55 二硫键对 hNgb 的氧化还原性质和 pH 依赖构象平衡的影响。我们发现 SS 桥显著影响血红素 Fe(III)到 Fe(II)还原焓（ΔH°'）和熵（ΔS°'），主要是由于还原诱导的溶剂重排效应的变化，而不影响轴向配体结合相互作用以及血红素环境的极性和静电。在 pH3 到 12 之间，高铁 hNgb 的血红素的电子性质对五个酸碱平衡敏感，而这些平衡几乎不受 Cys46/Cys55 二硫键的影响。极端 pH 值下发生的平衡诱导血红素释放，而在 pH5 到 10 之间发生的平衡则在不改变其轴向配位和低自旋状态的情况下改变血红素的电子性质。它们涉及靠近血红素的非配位氨基酸的侧链和至少一个血红素丙酸盐。
