Femtosecond Coherence Spectroscopy & Ultrafast Kinetic Study of Heme Proteins

飞秒相干光谱

• 批准号: 8209099
• 负责人: Paul M. Champion
• 金额: $ 32.49万
• 依托单位: NORTHEASTERN UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1984
• 资助国家: 美国
• 起止时间: 1984-06-01 至 2013-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8209099
• 关键词: Affect; Architecture; Basic Science; Behavior; Binding; Biochemical Reaction; Biological Models; Biological Process; Camphor 5-Monooxygenase; Cell Nucleus; Charge; Chemistry; Cytochrome P450; DNA Sequence Rearrangement; Discrimination; Distal; Docking; Electron Transport; Electronics; Entropy; Environment; Equilibrium; Event; Exhibits; Frequencies; Genetic Recombination; Goals; Grant; Health; Heme; Heme Group; Heme Iron; Hemeproteins; Human body; Investigation; Iron; Iron-Sulfur Proteins; Kinetics; Lead; Learning; Length; Ligand Binding; Ligands; Measures; Mediating; Metabolic Diseases; Metalloproteins; Metals; Methodology; Methods; Modeling; Molecular; Monitor; Motion; Myoglobin; Neutrons; Nitric Oxide Synthase; Nitrogen; Nuclear; Optics; Oxidation-Reduction; Oxygen; Peroxidases; Phase; Physiological; Play; Positioning Attribute; Process; Production; Protein S; Proteins; Pump; Pyrroles; Reaction; Research; Role; Signal Transduction; Soluble Guanylate Cyclase; Solutions; Specificity; Spectrum Analysis; Stretching; Structure; Sulfur; System; Techniques; Temperature; Testing; Time; Translating; Transport Process; Water; Work; absorption; aqueous; biological systems; chemical reaction; cofactor; cytochrome c; density; falls; insight; nitrophorin; protein complex; protein function; vibration

This project aims to extend our understanding of the structure, function, and dynamics of heme proteins such as cytochrome P450, mammalian peroxidases, nitric oxide synthase, soluble guanylate cyclase, nitrophorin, and cytochrome c. These proteins are involved in broad array of catalytic, signaling, and electron transport processes and are capable of an amazingly broad range of functions, even when the heme axial ligands are identical. This indicates that the protein architecture, and its influence on the heme structure, plays an important functional role. By using coherence spectroscopy, a femtosecond optical "pump-probe" technique, the "soft" out-of-plane (OOP) low-frequency vibrational modes of the heme can be excited and analyzed even in an aqueous environment. These vibrational modes have not been documented previously because they are difficult to access using traditional spectroscopic methods. They fall in the region of ambient thermal excitations (<200cm-1 ~300K) and are therefore most likely to be utilized as reaction coordinates by proteins. The observed coherence spectral intensities of these "soft" modes depend upon the magnitude of the heme structural distortions that are induced by the protein architecture. These OOP heme motions are functionally significant, as demonstrated by the importance of the heme "doming" mode in the diatomic ligand binding reaction. The rich spectrum of the low-frequency heme motions is just beginning to be appreciated, as a wider variety of proteins and model compounds is examined. This project aims to explore the functional role of both static distortions and thermally excited low-frequency vibrations in heme proteins. Distortions (such as heme "ruffling" and "saddling") that alter the electronic orbital interactions between the iron atom and its surrounding molecular framework are hypothesized to affect the redox potential of the metal center. Vibrational motions along these same, thermally accessible, OOP coordinates are excellent candidates to mediate and control electron transfer. Coherence spectroscopy is uniquely positioned to probe these modes in aqueous solution. For example, we will examine electron transfer partners, such as Pdx and CYP101, in order to monitor changes in the low frequency spectrum that occur when the protein complex is formed. The low frequency modes of Fe-S proteins will also be examined. Kinetic probes on ultrafast timescales, stretching over 10 decades in time, will be used to study the rapid time-scale, non-equilibrium processes, that take place immediately following the electronic rearrangements associated with biochemical reactions. For example, the two geminate phases for oxygen rebinding to the heme in myoglobin exhibit very different Arrhenius prefactors, suggesting that the entropic barrier for recombination is time dependent. Such non-equilibrium processes will be studied to learn if they allow heme proteins to enhance discrimination between different classes of diatomic ligands.

这个项目旨在扩展我们对血红素蛋白的结构、功能和动力学的理解 如细胞色素P450、哺乳动物过氧化物酶、一氧化氮合酶、可溶性鸟苷酸环化酶 硝基转运蛋白和细胞色素C。这些蛋白质参与了广泛的催化、信号传导和 电子传输过程，并能够一个惊人的广泛的功能，即使当 血红素轴向配体是相同的。这表明，蛋白质结构及其对蛋白质结构的影响， 血红素结构，起着重要的功能作用。通过使用相干光谱学， 光学“泵浦-探测”技术，“软”出平面（OOP）的低频振动模式的 血红素甚至可以在水性环境中被激发和分析。这些振动模式没有 因为它们很难使用传统的光谱技术进行访问， 方法.它们属于环境热激发区（<200 cm-1 ~ 300 K），因此是最重要的 可能被蛋白质用作反应坐标。观察到的相干光谱强度 这些“软”模式取决于由 蛋白质结构。这些OOP血红素运动在功能上是重要的，如图所示。 血红素“圆顶”模式在双原子配体结合反应中的重要性。丰富的光谱 低频血红素运动才刚刚开始受到重视，因为更广泛的蛋白质和模型 化合物进行了检查。本项目旨在探讨静态扭曲和 血红素蛋白质中的热激发低频振动。扭曲（如血红素“起皱”和 “鞍”），改变铁原子与其周围环境之间的电子轨道相互作用 假设分子骨架影响金属中心的氧化还原电位。振动 沿着这些相同的、热可达的OOP坐标的运动是调解的最佳候选者 并控制电子转移。相干光谱是唯一定位于探测这些模式， 水溶液中例如，我们将研究电子转移伙伴，如Pdx和CYP 101， 以便监测蛋白质复合物形成时发生的低频频谱的变化。 还将检查Fe-S蛋白的低频模式。超快时间尺度上的动力学探测器， 将被用来研究快速的时间尺度，非平衡过程， 在与生物化学相关的电子重排之后立即发生， 反应.例如，氧与肌红蛋白中的血红素重新结合的两个成对相表现出 非常不同的Arabius前因子，这表明重组的熵障碍是时间 依赖。这种非平衡过程将被研究，以了解它们是否允许血红素蛋白质 增强不同类别的双原子配体之间的区分。

项目成果

Investigations of heme ligation and ligand switching in cytochromes p450 and p420.

• DOI: 10.1021/bi400541v
• 发表时间: 2013-08-27
• 期刊: BIOCHEMISTRY
• 影响因子: 2.9
• 作者: Sun, Yuhan;Zeng, Weiqiao;Benabbas, Abdelkrim;Ye, Xin;Denisov, Ilia;Sligar, Stephen G.;Du, Jing;Dawson, John H.;Champion, Paul M.
• 通讯作者: Champion, Paul M.

Investigations of ferric heme cyanide photodissociation in myoglobin and horseradish peroxidase.

肌红蛋白和辣根过氧化物酶中血红素氰化铁光解的研究。

• DOI: 10.1021/jp401224f
• 发表时间: 2013
• 期刊: The journal of physical chemistry. B
• 作者: Zeng,Weiqiao;Sun,Yuhan;Benabbas,Abdelkrim;Champion,PaulM
• 通讯作者: Champion,PaulM

Investigations of the low frequency modes of ferric cytochrome c using vibrational coherence spectroscopy.

• DOI: 10.1021/jp501298c
• 发表时间: 2014-06-12
• 期刊: The journal of physical chemistry. B
• 作者: Karunakaran V;Sun Y;Benabbas A;Champion PM
• 通讯作者: Champion PM

Investigations of the low-frequency spectral density of cytochrome c upon equilibrium unfolding.

• DOI: 10.1021/jp404881k
• 发表时间: 2013-08-22
• 期刊: The journal of physical chemistry. B
• 作者: Sun Y;Karunakaran V;Champion PM
• 通讯作者: Champion PM

Temperature-dependent heme kinetics with nonexponential binding and barrier relaxation in the absence of protein conformational substates.

在不存在蛋白质构象亚状态的情况下，温度依赖性血红素动力学具有非指数结合和屏障松弛。

• DOI: 10.1073/pnas.0702622104
• 发表时间: 2007
• 期刊: Proceedings of the National Academy of Sciences of the United States of America
• 影响因子: 11.1
• 作者: Ye,Xiong;Ionascu,Dan;Gruia,Florin;Yu,Anchi;Benabbas,Abdelkrim;Champion,PaulM
• 通讯作者: Champion,PaulM