Du P, Loew G H
Molecular Research Institute, Palo Alto, California 94304.
Biophys J. 1995 Jan;68(1):69-80. doi: 10.1016/S0006-3495(95)80160-8.
Theoretical studies of the electronic structure and spectra of models for the ferric resting state and Compound I intermediates of horseradish peroxidase (HRP-I) and catalase (CAT-I) have been performed using the INDO-RHF/CI method. The goals of these studies were twofold: i) to determine whether the axial ligand of HRP is best described as imidazole or imidazolate, and ii) to address the long-standing question of whether HRP-I and CAT-I are a1u and a2u tau cation radicals. Only the imidazolate HRP-I model led to a calculated electronic spectra consistent with the experimentally observed significant reduction in the intensity of the Soret band compared with the ferric resting state. These results provide compelling evidence for significant proton transfer to the conserved Asp residue by the proximal histidine. The origin of the observed reduction of the Soret band intensity in HRP-I and CAT-I spectra has been examined and found to be caused by the mixing of charge transfer transitions into the predominantly porphyrin tau-tau transitions. For both HRP-I and CAT-I, the a1u porphyrin tau cation state is the lowest energy, and it is further stabilized by both the anionic form of the ligand and the porphyrin ring substituents of protoporphyrin-IX. The calculated values of quadrupole-splitting observed in the Mossbauer resonance of HRP-I and CAT-I are similar for the a1u and a2u tau cation radicals. Electronic spectrum of the a1u tau cation radical of HRP-I are more similar to the observed spectra, whereas the spectra of both a1u tau and a2u tau cation radicals of CAT-I resemble the observed spectra. These results also indicate the limitations of using any one observable property to try to distinguish between these states. Taken together, comparison of calculated and observed properties indicate that there is no compelling reason to invoke the higher energy a2u tau cation radical as the favored state in HRP-I and CAT-I. Both ground-state properties and electronic spectra are consistent with the a1u tau cation radical.
采用INDO - RHF/CI方法对辣根过氧化物酶（HRP - I）和过氧化氢酶（CAT - I）的铁静止态和化合物I中间体模型的电子结构和光谱进行了理论研究。这些研究的目标有两个：i）确定HRP的轴向配体最好描述为咪唑还是咪唑盐，ii）解决长期存在的问题，即HRP - I和CAT - I是否为a1u和a2u τ阳离子自由基。只有咪唑盐HRP - I模型得出的计算电子光谱与实验观察到的与铁静止态相比Soret带强度显著降低一致。这些结果为近端组氨酸向保守的天冬氨酸残基发生显著质子转移提供了有力证据。已对HRP - I和CAT - I光谱中观察到的Soret带强度降低的起源进行了研究，发现这是由于电荷转移跃迁混入主要的卟啉τ - τ跃迁所致。对于HRP - I和CAT - I，a1u卟啉τ阳离子态能量最低，并且通过配体的阴离子形式和原卟啉 - IX的卟啉环取代基进一步稳定。在HRP - I和CAT - I的穆斯堡尔共振中观察到的四极分裂计算值对于a1u和a2u τ阳离子自由基相似。HRP - I的a1u τ阳离子自由基的电子光谱与观察到的光谱更相似，而CAT - I的a1u τ和a2u τ阳离子自由基的光谱都类似于观察到的光谱。这些结果还表明仅使用任何一种可观测性质来区分这些状态的局限性。综合来看，计算性质与观察性质的比较表明，没有令人信服的理由将能量更高的a2u τ阳离子自由基作为HRP - I和CAT - I中的优势态。基态性质和电子光谱都与a1u τ阳离子自由基一致。
