Near Ultraviolet Raman Studies of Cytochrome P450

描述（由申请人提供）：我们的目标是通过使用泵浦-探测动力学和时间分辨共振拉曼光谱在宽的动态时间范围（ps-ms）内扩展我们对细胞色素P450和其他血红素蛋白系统的结构和功能的理解。一个独特的时间稳定，电子可调谐，双色皮秒激光系统的发展将使瞬态拉曼和动力学研究。飞秒相干光谱（FCS），这是一个直接的和独特的探针的相干蛋白质/辅因子的振动运动诱导的电子跃迁与生化反应，也将被用于研究的生化反应坐标以及全球蛋白质响应。一个具体的假设进行测试地址背后的原因，非常低的飞秒量子产率O2光解和潜在的瞬态侧对氧结合的几何形状，因为配体弯曲模式的参与解离的反应坐标中出现。振动热六坐标氧合肌红蛋白物种已经被检测到使用白色光连续探针，我们建议将这样的研究扩展到P450系统及其催化中间体，以及扩展探测能力，包括皮秒拉曼散射技术。还将利用新开发的光学技术进行低温实验。这些实验将包括第一次皮秒和飞秒的研究作为温度的函数的双生配体重组。探索工作与拉曼显微镜将继续与线粒体的空间特异性探针和嵌入在膜纳米盘的P450的研究。 总体而言，该项目具有广泛的健康相关影响，特别是涉及细胞色素P450和血红素蛋白。许多化疗药物以及多环致癌物都是通过P450系统代谢的。此外，所有涉及P450蛋白的代谢紊乱都必须与本研究有关，因为在分子水平上对P450活性位点结构和功能的基本理解将使那些关心这些疾病治疗的人更好地了解任何水平。

DESCRIPTION (provided by applicant): We aim to extend our understanding of the structure and function of cytochrome P450 and other heme protein systems by using pump-probe kinetics and time resolved resonance Raman spectroscopy over a wide dynamic range in time (ps-ms). Development of a unique temporally stabilized, and electronically tunable, two-color picosecond laser system will enable both the transient Raman and the kinetic studies. Femtosecond coherence spectroscopy (FCS), which is a direct and unique probe of the coherent protein/cofactor vibrational motion induced by electronic transitions associated with biochemical reactions, will also be utilized in studies of the biochemical reaction coordinates as well as the global protein response. One specific hypothesis to be tested addresses the reason behind the anomalously low femtosecond quantum yield for O2 photolysis and the potential for a transient side-on oxygen binding geometry that arises because of the participation of ligand bending modes in the reaction coordinate for dissociation. Vibrationally hot six coordinate oxymyoglobin species have already been detected using white light continuum probes, and we propose to extend such studies to the P450 system and its catalytic intermediates as well as to expand the probe capability to include picosecond Raman scattering techniques. Experiments at low temperature will also be carried out using newly developed optical techniques. These experiments will include the first picosecond and femtosecond studies of geminate ligand recombination as a function of temperature. Exploratory work with the Raman microscope will continue with spatially specific probes of mitochondria and with studies of P450 embedded in membrane nanodiscs. Overall this project has a wide range of health related implications involving cytochrome P450 in particular and heme proteins in general. Many chemotherapeutic agents as well as polycyclic carcinogens are metabolized by P450 systems. Moreover, all metabolic disorders involving a P450 protein must relate to this research, since a fundamental understanding of the active site structure and function of P450 at the molecular level will result in a better insight for those concerned with treatment of these disorders at any level.