The H93G myoglobin cavity mutant as a versatile template for modeling heme proteins: ferrous, ferric, and ferryl mixed-ligand complexes with imidazole in the cavity.
The H93G myoglobin cavity mutant as a versatile template for modeling heme proteins: ferrous, ferric, and ferryl mixed-ligand complexes with imidazole in the cavity.
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DOI：
10.1021/ic0007198
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发表时间：
2000-12
影响因子：
4.6
通讯作者：
A. Pond;M. Roach;M. R. Thomas;S. Boxer;J. Dawson
中科院分区：
文献类型：
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作者：
A. Pond;M. Roach;M. R. Thomas;S. Boxer;J. Dawson
制备血红素蛋白精确的室温模型复合物的困难之一，特别是在铁态下，是混合配体加合物的产生：血红素两侧具有不同配体的复合物。肌红蛋白（Mb） H93G空腔突变体血红素两侧可及性的差异为这一问题提供了一个潜在的通用解决方案。为了证明H93G - Mb在制备血红素蛋白模型中的通用性，已经制备了许多亚铁、铁和铁基咪唑连接的H93G（H93G(Im) Mb）的混合配体加合物。与类似的野生型Mb衍生物相比，配合物的电子吸收和磁性圆二色性（MCD）光谱进行了表征。起始铁H93G(Im) Mb状态在光谱上与野生型Mb铁相似，这与在近端腔中含有单个咪唑而在远端含有水键的配合物的预期相似。在铁H93G(Im) Mb上加入第六配体，无论是电荷中性的（咪唑）还是阴离子的（氰化物和叠氮化物），都会形成六配位低自旋配合物，其MCD特征与野生型铁Mb的平行衍生物相似。还原铁H93G(Im) Mb并随后暴露于CO、NO或O2中产生铁配合物(脱氧、CO、NO、和O2)的MCD光谱一直与野生型亚铁Mb的类似亚铁物种相似。H93G(Im) Mb铁与H2O2反应形成稳定的高价氧化铁基复合物，具有MCD特征，与氧化铁基野生型Mb基本相同。如此广泛的混合配体血红素复合物的产生证明了H93G Mb空腔突变体作为制备血红素蛋白模型复合物模板的有效性。
One of the difficulties in preparing accurate ambient-temperature model complexes for heme proteins, particularly in the ferric state, has been the generation of mixed-ligand adducts: complexes with different ligands on either side of the heme. The difference in the accessibility of the two sides of the heme in the H93G cavity mutant of myoglobin (Mb) provides a potential general solution to this problem. To demonstrate the versatility of H93G Mb for the preparation of heme protein models, numerous mixed-ligand adducts of ferrous, ferric, and ferryl imidazole-ligated H93G (H93G(Im) Mb) have been prepared. The complexes have been characterized by electronic absorption and magnetic circular dichroism (MCD) spectroscopy in comparison to analogous derivatives of wild type Mb. The starting ferric H93G(Im) Mb state spectroscopically resembles wild-type ferric Mb as expected for a complex containing a single imidazole in the proximal cavity and water bound on the distal side. Addition of a sixth ligand to ferric H93G(Im) Mb, whether charge neutral (imidazole) or anionic (cyanide and azide), results in formation of six-coordinate low-spin complexes with MCD characteristics similar to those of parallel derivatives of wild-type ferric Mb. Reduction of ferric H93G(Im) Mb and subsequent exposure to either CO, NO, or O2 produces ferrous complexes (deoxy, CO, NO, and O2) that consistently exhibit MCD spectra similar to the analogous ferrous species of wild-type ferrous Mb. Most interestingly, reaction of ferric H93G(Im) Mb with H2O2 results in the formation of a stable high-valent oxoferryl complex with MCD characteristics that are essentially identical to those of oxoferryl wild-type Mb. The generation of such a wide array of mixed-ligand heme complexes demonstrates the efficacy of the H93G Mb cavity mutant as a template for the preparation of heme protein model complexes.