Mono- and bis-phosphine-ligated H93G myoglobin: spectral models for ferrous-phosphine and ferrous-CO cytochrome P450.
Mono- and bis-phosphine-ligated H93G myoglobin: spectral models for ferrous-phosphine and ferrous-CO cytochrome P450.
复制标题
单膦和双膦连接的 H93G 肌红蛋白：亚铁膦和亚铁 CO 细胞色素 P450 的光谱模型。
DOI：
10.1016/j.jinorgbio.2013.03.006
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发表时间：
2013
影响因子：
3.9
通讯作者：
Dawson,JohnH
中科院分区：
文献类型：
--
作者：
Sun,Shengfang;Sono,Masanori;Dawson,JohnH
为了进一步研究磷化氢作为血红素蛋白结构探针和功能探针的性质，分别用THMP滴定外源无配体铁和亚铁H93G Mb，制备了H93G肌红蛋白（Mb）的单和双磷化氢[三（羟甲基）磷化氢，THMP]加合物。用CO泡泡或咪唑（Im）逐步滴定双THMP连接的亚铁蛋白生成混合配体（分别为THMP/CO或THMP/Im）亚铁配合物。将单THMP- fe （II）蛋白用O2鼓泡，在−40℃下形成稳定的氧化亚铁H93G(THMP) Mb。在单thmp加合物中加入h2o2后，thmp连接的铁基H93G Mb部分形成。本研究用紫外可见吸收光谱和磁圆二色光谱对所制备的物质进行了表征。H93G Mb的六配位亚铁双膦和单膦/CO配合物表现出独特的光谱特征（分别为红移Soret/独特形状的MCD可见光谱和高卟啉光谱），这些特征只有在硫代酸连接的血红素蛋白（如细胞色素P450 （P450）和富烟氯过氧化物caldariomyces fumagochloroperoxidase (CPO)）的类似膦或CO配合物中才能看到。然而，在磷化氢结合的铁H93G Mb中没有这种相似性，尽管磷化氢结合的铁P450和CPO显示高卟啉光谱。事实上，双thmp结合的铁H93G Mb具有与双胺和双硫醚连接的H93G Mb配合物相似的MCD和紫外-可见吸收光谱。本研究还进一步证明了H93G空腔突变体在制备新型血红素铁配位结构中的实用性。
To further investigate the properties of phosphines as structural and functional probes of heme proteins, mono- and bis-phosphine [tris(hydroxymethyl)phosphine, THMP] adducts of H93G myoglobin (Mb) have been prepared by stepwise THMP titrations of exogenous ligand-free ferric and ferrous H93G Mb, respectively. Bubbling with CO or stepwise titration with imidazole (Im) of the bis-THMP-ligated ferrous protein generated a mixed ligand (THMP/CO or THMP/Im, respectively) ferrous complexes. Stable oxyferrous H93G(THMP) Mb was formed at − 40 °C by bubbling the mono-THMP-Fe(II) protein with O2. A THMP-ligated ferryl H93G Mb moiety has been partially formed upon addition of H2O2to the ferric mono-THMP adduct. All the species prepared above have been characterized with UV–visible (UV–vis) absorption and magnetic circular dichroism (MCD) spectroscopy in this study. The six-coordinate ferrous bis-phosphine and mono-phosphine/CO complexes of H93G Mb exhibit characteristic spectral features (red-shifted Soret/unique-shaped MCD visible bands and hyperporphyrin spectra, respectively) that only have been seen for the analogous phosphine or CO-complexes of thiolate-ligated heme proteins such as cytochrome P450 (P450) andCaldariomyces fumagochloroperoxidase (CPO). However, such resemblance is not seen in phosphine-ligated ferric H93G Mb even though phosphine-bound ferric P450 and CPO display hyperporphyrin spectra. In fact, bis-THMP-bound ferric H93G Mb exhibits MCD and UV–vis absorption spectra that are similar to those of bis-amine- and bis-thioether-ligated H93G Mb complexes. This study also further demonstrates the utility of the H93G cavity mutant for preparing novel heme iron coordination structures.