International Collaboration in Chemistry: Protein Dynamics and Heme Protein Function

将详细研究人吲哚2，3-双加氧酶和一氧化氮合酶催化的生理过程，包括双原子配体(O2，NO，CO)、底物和血红素铁之间的反应，以获得对反应机理的原子水平描述。这种方法包括对蛋白质结构、结构波动和松弛的分析，以及蛋白质内配体和底物的动力学。我们将使用稳态和时间分辨光谱技术，并在光谱的红外和紫外/可见光区域进行监测。配体将被激光脉冲光解，以及随后的过程(配体结合、底物结合、蛋白质松弛等)。将在很大的时间和温度范围内(3-300K)观察到。低温下的测量使我们能够制备和保存稀有和短寿命的反应中间体，随后可以对其进行详细分析。这些方法是近年来以肌红蛋白为模型系统发展起来的，并已成功应用于其他血红素蛋白。将特别强调生理上重要的配体NO的反应。为了评估特定氨基酸对能量格局的影响，从而对活性部位的性质以及蛋白质内的配体迁移和与蛋白质结合的底物的影响，蛋白质将通过定点突变进行修饰。

The physiological processes catalyzed by human indole 2,3-dioxygenase and nitric oxide synthase, which involve the reaction between a diatomic ligand (O2, NO, CO), a substrate and the heme iron will be studied in detail to obtain an atomic-level description of the reaction mechanisms. This approach involves the analysis of protein structures, structural fluctuations and relaxations, and the dynamics of ligands and substrates within the protein. We will employ steady-state and time-resolved spectroscopic techniques with monitoring in the infrared and UV/visible regions of the spectrum. Ligands will be photodissociated by laser pulses, and the ensuing processes (ligand binding, substrate binding, protein relaxations etc.) will be observed over wide ranges of time and temperature (3 – 300 K). Measurements at cryogenic temperatures allow us to prepare and conserve rare and short-lived reaction intermediates, which can subsequently be analyzed in detail. These methods have been developed in recent years with myoglobin as a model system and have already been successfully applied to other heme proteins. Special emphasis will be put on reactions of the physiologically important ligand NO. To assess the influence of specific amino acids on the energy landscape and thus on active-site properties and ligand migration within and substrate binding to the protein, the proteins will be modified by site-directed mutagenesis.