Modeling the active site of cytochrorne oxidase:: Synthesis and characterization of a cross-linked histidine-phenol
Modeling the active site of cytochrorne oxidase:: Synthesis and characterization of a cross-linked histidine-phenol
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DOI：
10.1021/ja011852h
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发表时间：
2002-02-27
影响因子：
15
通讯作者：
Einarsdóttir, O
中科院分区：
文献类型：
--
作者：
Cappuccio, JA;Ayala, I;Einarsdóttir, O
合成了交联组氨酸苯酚化合物作为细胞色素 c 氧化酶活性位点的化学类似物。通过IR、H-1和C-13 NMR、质谱和单晶X射线分析验证了交联化合物（化合物1）的结构。分光光度滴定表明化合物 1 上酚质子的 pK(a) (8.34) 低于酪氨酸 (10.1) 或对甲酚 (10.2) 的 pK(a)。 pK(a) 的这种降低与交联的组氨酸-酪氨酸可能促进质子递送至细胞色素 c 氧化酶中的双核位点的假设一致。化合物 1 在室温下的时间分辨光学吸收光谱（在存在和不存在分子氧的情况下在 266 nm 处激发）表明在类似于 330 和类似于 500 nm 处具有最大吸收的物质，我们将其指定为化合物 1 的苯氧基。在 UV 光解后获得的化合物 1 的电子顺磁共振 (EPR) 光谱证实了 低温下的顺磁性物质。由于交联化合物缺乏 β-亚甲基质子，因此与酪氨酰自由基相比，EPR 线形状发生了显着改变。然而，EPR 线形状的模拟和各向同性 g 值的测量与咪唑氮的少量耦合以及苯氧基环中的自旋密度扰动很小是一致的。化合物1的基态傅里叶变换红外(FT-IR)光谱表明，咪唑环的添加扰乱了酪氨酸环伸缩振动的频率。与交联化合物的氧化相关的差异 FT-IR 光谱检测到苯氧基自由基振动带的显着扰动。我们假设苯酚氧化会在化合物 1 的咪唑氮上产生较小的自旋密度离域，这可以解释其独特的光谱特性。
A cross-linked histidine-phenol compound was synthesized as a chemical analogue of the active site of cytochrome c oxidase. The structure of the cross-linked compound (compound 1) was verified by IR, H-1 and C-13 NMR, mass spectrometry, and single-crystal X-ray analysis. Spectrophotometric titrations indicated that the pK(a) of the phenolic proton on compound 1 (8.34) was lower than the pK(a) of tyrosine (10.1) or of p-cresol (10.2). This decrease in pK(a) is consistent with the hypothesis that a cross-linked histidine-tyrosine may facilitate proton delivery to the binuclear site in cytochrome c oxidase. Time-resolved optical absorption spectra of compound 1 at room temperature, generated by excitation at 266 nm in the presence and absence of dioxygen, indicated a species with absorption maxima at similar to330 and similar to500 nm, which we assign to the phenoxyl radical of compound 1. The electron paramagnetic resonance (EPR) spectra of compound 1, obtained after UV photolysis, confirmed the generation of a paramagnetic species at low temperature. Because the cross-linked compound lacks beta-methylene protons, the EPR line shape was dramatically altered when compared to that of the tyrosyl radical. However, simulation of the EPR line shape and measurement of the isotropic g value was consistent with a small coupling to the imidazole nitrogen and with little spin density perturbation in the phenoxyl ring. The ground-state Fourier transform infrared (FT-IR) spectrum of compound 1 showed that addition of the imidazole ring perturbs the frequency of the tyrosine ring stretching vibrations. The difference FT-IR spectrum, associated with the oxidation of the cross-linked compound, detected significant perturbations of the phenoxyl radical vibrational bands. We postulate that phenol oxidation produces a small delocalization of spin density onto the imidazole nitrogen of compound 1, which may explain its unique optical spectral properties.