Investigations of Low-Frequency Vibrational Dynamics and Ligand Binding Kinetics of Cystathionine β-Synthase
Investigations of Low-Frequency Vibrational Dynamics and Ligand Binding Kinetics of Cystathionine β-Synthase
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DOI：
10.1021/jp909700r
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发表时间：
2010-03-11
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;Benabbas, Abdelkrim;Champion, Paul M.
利用振动相干光谱研究了人胱硫醚β-合酶（CBS）截短二聚体的低频动力学。CBS是具有半胱氨酸和组氨酸轴向配体的吡哆醛-5 '-磷酸依赖性血红素酶，其催化丝氨酸和高半胱氨酸缩合形成胱硫醚。“失谐”相干谱（探测更高的频率）和拉曼谱之间的强相关性被证明，并揭示了低于200 cm（-1）的模式丰富的图案。铁CBS晶体结构的正常坐标结构分解（NSD）预测了具有显著血红素“隆起”、“褶皱”和“鞍形”含量的正常模式的增强，并且它们在接近40、接近60和接近90 cm（-1）的相干光谱中观察到。当pH变化时，低频血红素模式的相对强度和频率表明在pH 7附近存在其独特的蛋白质诱导的血红素结构扰动，这与在更高或更低pH下观察到的不同。对于铁CBS，我们观察到一个新的模式，接近25 cm（-1），可能涉及蛋白质的响应，其对于在Soret手最大值的蓝色和红色侧上的激发表现出类似于π的相位跳变。沿着我们对亚铁CBS的NO键合配合物的探测，也给出了亚铁CBS的低频振动相干谱。CBS的CO双生再结合动力学与基因激活蛋白CooA的CO结合形式相似，但出现了显著的额外动力学不均匀性。对这种不均匀性的分析表明，它是由CBS的两个亚基引起的，并导致两个亚基的平均CO成对重结合率的比值的近似因子为20。
Vibrational coherence spectroscopy is used to study the low frequency dynamics of the truncated dimer of human cystathionine beta-synthase (CBS). CBS is it pyridoxal-5'-phosphate-dependent heme enzyme with cysteine and histidine axial ligands that catalyzes the condensation of serine and homocysteine to form cystathionine. A strong correlation between the "detuned" coherence spectrum (which probes higher frequencies) and the Raman spectrum is demonstrated, and a rich pattern of modes below 200 cm(-1) is revealed. Normal coordinate structural decomposition (NSD) of the ferric CBS crystal structure predicts the enhancement of normal modes with significant heme "doming", "ruffling", and "saddling" content, and they are observed in the coherence spectra near similar to 40, similar to 60, and similar to 90 cm(-1). When pH is varied, the relative intensities and frequencies of the low frequency heme modes indicate the presence of it unique protein-induced heme structural perturbation near pH 7 that differs from what is observed at higher or lower pH. For ferric CBS, we observe a new mode near similar to 25 cm(-1), possibly involving the response of the protein, which exhibits a phase jump of similar to pi for excitation on the blue and red side of the Soret hand maximum. The low frequency vibrational coherence spectrum of ferrous CBS is also presented, along with Our efforts to probe its NO-bound complex. The CO geminate rebinding kinetics of CBS are similar to the CO-bound form of the gene activator protein CooA, but with the appearance of a significant additional kinetic inhomogeneity. Analysis of this inhomogeneity suggests that it arises from the two Subunits of CBS and leads to it factor of similar to 20 for the ratio of the average CO geminate rebinding rates of the two subunits.