Investigation of the low frequency dynamics of heme proteins: native and mutant cytochrome P450(cam) and redox partner complexes.
Investigation of the low frequency dynamics of heme proteins: native and mutant cytochrome P450(cam) and redox partner complexes.
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DOI：
10.1021/jp112298y
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发表时间：
2011-05-12
影响因子：
3.3
通讯作者：
Champion, Paul M.
中科院分区：
文献类型：
--
作者：
Karunakaran, Venugopal;Denisov, Ilia;Sligar, Stephen G.;Champion, Paul M.
振动相干光谱 (VCS) 用于研究不含樟脑和结合樟脑的细胞色素 P450cam (CYP 101) 及其 L358P 突变体的低频动力学。发现低频血红素振动在与电子转移伙伴恶臭氧还蛋白 (Pdx) 结合时受到干扰。还证明了“失谐”振动相干谱（监测 100-400 cm−1 之间的频率）与拉曼谱的较低频率部分之间的强相关性。极低频区域 ≤ 200 cm−1，通过开带 VCS 测量唯一访问，揭示了当远端口袋中不存在樟脑时，P450cam 中接近 103 cm−1 的模式。这反映了在水与低自旋铁原子配位的无底物状态下存在特定的血红素扭曲，例如鞍状或皱褶。这种扭曲可能会降低电子转移到无底物蛋白质的速率。樟脑束缚形式中约 33 cm−1 附近强模式的存在表明存在显着的血红素圆顶变形，使用正态坐标结构分解的分析支持了这一点。 Pdx 还在 30 cm−1 附近表现出强烈的相干振动，原则上可能参与与其电子转移目标的振动共振。当 P450cam/Pdx 复合体形成时，会出现 33 cm−1 特征的分裂和 78 cm−1 附近模式的强化。这些观察结果与 Pdx 结合时的振动混合和血红素几何变形一致，这与向血红素提供的硫醇盐电子增加一致。 L358P 突变体的相干光谱中 65 cm−1 附近模式的出现与 P450cam/Pdx 复合物中看到的 78 cm−1 模式相当，并且与 L358P 突变体中的血红素及其环境与 Pdx 结合的天然蛋白相似的观点一致。显示了 P450cam 和 L358P 突变体的共振拉曼光谱，这些变化与突变体样品中向血红素提供硫醇盐电子的增加量相关。
Vibrational coherence spectroscopy (VCS) is used to investigate the low frequency dynamics of camphor-free and camphor-bound cytochrome P450cam (CYP 101) and its L358P mutant. The low frequency heme vibrations are found to be perturbed upon binding to the electron transfer partner putidaredoxin (Pdx). A strong correlation between the “detuned” vibrational coherence spectrum, which monitors frequencies between 100–400 cm−1, and the lower frequency part of the Raman spectrum is also demonstrated. The very low frequency region ≤ 200 cm−1, uniquely accessed by open-band VCS measurements, reveals a mode near 103 cm−1 in P450cam when camphor is not present in the distal pocket. This reflects the presence of a specific heme distortion, such as saddling or ruffling, in the substrate-free state where water is coordinated to the low-spin iron atom. Such distortions are likely to retard the rate of electron transfer to the substrate-free protein. The presence of strong mode near ~33 cm−1 in the camphor bound form suggests a significant heme doming distortion, which is supported by analysis using normal coordinate structural decomposition. Pdx also displays a strong coherent vibration near 30 cm−1 that could, in principle, be involved in vibrational resonance with its electron transfer target. A splitting of the 33 cm−1 feature and intensification of a mode near 78 cm−1 appear when the P450cam/Pdx complex is formed. These observations are consistent with vibrational mixing and heme geometric distortions upon Pdx binding that are coincident with the increased thiolate electron donation to the heme. The appearance of a mode near 65 cm−1 in the coherence spectra of the L358P mutant is comparable to the mode at 78 cm−1 seen in the P450cam/Pdx complex and is consistent with the view that the heme and its environment in the L358P mutant are similar to the Pdx-bound native protein. Resonance Raman spectra are presented for both P450cam and the L358P mutant and the changes are correlated with an increased amount of thiolate electron donation to the heme in the mutant sample.
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影响因子：
3.3
作者：
Karunakaran, Venugopal;Benabbas, Abdelkrim;Champion, Paul M.
通讯作者：
Champion, Paul M.
影响因子：
2.9
作者：
Ma, JG;Zhang, J;Shelnutt, JA
通讯作者：
Shelnutt, JA
影响因子：
2.9
作者：
Gruia, Flavin;Ionascu, Dan;Champion, Paul M.
通讯作者：
Champion, Paul M.
影响因子：
3.4
作者：
Gruia, Flaviu;Ye, Xiong;Champion, Paul M.
通讯作者：
Champion, Paul M.
影响因子：
15
作者：
Gruia, Flaviu;Kubo, Minoru;Champion, Paul M.
通讯作者：
Champion, Paul M.