Ultrafast Spectroscopic Methods to Probe Photodamage and Unfolding in Biopolymers

超快光谱方法探测生物聚合物中的光损伤和展开

• 批准号: 7761111
• 负责人: David W McCamant
• 金额: $ 19.13万
• 依托单位: UNIVERSITY OF ROCHESTER
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-08-01 至 2013-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7761111
• 关键词: Address; Amino Acids; Area; Aromatic Amino Acids; Biochemical Process; Biology; Biopolymers; Characteristics; Collaborations; Complement; DNA; DNA lesion; Data; Dependence; Development; Dimerization; Dissociation; Electronics; Environment; Event; Evolution; Exhibits; Fluorescence; Fostering; Funding; Goals; Health; Heating; Human; Human Biology; Investigation; Kinetics; Link; Mechanics; Methodology; Methods; Mission; Modeling; Molecular Structure; Monitor; Motion; Nature; Nucleic Acids; Oligonucleotides; Optics; Outcome; Peptides; Photobiology; Photochemistry; Polymers; Positioning Attribute; Process; Production; Proteins; Public Health; Pyrimidine; Pyrimidines; Raman Spectrum Analysis; Research; Research Design; Research Personnel; Resolution; Side; Specificity; Spectrum Analysis; Structure; Structure-Activity Relationship; System; Techniques; Technology; Testing; Theoretical model; Time; UV induced; Ultraviolet Rays; United States National Institutes of Health; Universities; Work; absorption; aqueous; biological systems; environmental mutagens; innovation; instrument; instrumentation; interest; melting; molecular dynamics; nanosecond; novel therapeutics; protein function; public health relevance; quantum; quantum chemistry; tool; ultraviolet

DESCRIPTION (provided by applicant): New spectroscopic methods are needed to determine the structural changes that occur in biological systems on the femtosecond (10-15 s) to nanosecond (10-9 s) time-scale. Photodamage to nucleic acids by ultraviolet (UV) light occurs in 100's of femtoseconds and numerous functions of peptides, proteins and oligonucleotides depend on structural fluctuations occurring over picoseconds to nanoseconds and longer. In each of these cases, there are few experimental techniques that can resolve the dynamic molecular structure of the evolving system. Our work aims to correct this by developing ultrafast Raman spectroscopy capable of collecting vibrational spectra, which can be directly related to molecular structure, of photochemically and thermally activated dynamics in biomolecules. Specific Aim #1 of this proposal will develop new femtosecond Raman instrumentation that can collect high-resolution vibrational spectra with time resolution better than 100 fs. This methodology will be applied to gain new understanding of the ultrafast dimerization of pyrimidines in DNA following excitation by UV light, as well as new fundamental understanding of the quantum mechanical nature of the excitation in nucleic acid polymers. With Specific Aim #2, we will develop new methodologies to impulsively initiate thermal unfolding of biopolymers. This will allow the numerous tools of time-resolved spectroscopy, which currently have applications limited to photochemistry, to unravel the dynamics of thermally driven secondary structural changes on the picosecond to nanosecond time-scale. Raman spectroscopy is uniquely positioned to contribute to these areas because of its ability to collect vibrational spectra of biopolymers over a wide spectral window without interference from the aqueous environment. Raman spectra of biopolymers exhibit particular peaks that are characteristic of the secondary structure of the polymer and the particular environment of the side chains or nucleic acids. Hence by collecting time-resolved Raman spectra as biochemical processes proceed, we can determine both the time-scales of formation and the structures of kinetic intermediates. This experimental work will complement the many theoretical predictions that have been made about ultrafast structural changes in photoexcited DNA and longer time-scale structural fluctuations. The proposed research is significant because of the breadth of photobiology that can be addressed by these techniques and the need for experimental probes of rapid structural changes important to biology and human health. PUBLIC HEALTH RELEVANCE (provided by applicant): The proposed research directly supports the mission of the NIH by helping to establish new understanding of the mechanisms of ultraviolet light damage to DNA, and its implication for public health. Ultraviolet light is one of the most prevalent environmental mutagens on earth, with significant deleterious effects on human health. These studies will also increase the capability of biomedical researchers to investigate the rapid structural motions important to the functions of proteins and DNA, thereby accelerating the development of new therapeutics.

描述（由申请人提供）：需要新的光谱方法来确定生物系统中在飞秒（10-15 s）至纳秒（10-9 s）时间尺度上发生的结构变化。紫外（UV）光对核酸的光损伤发生在数百飞秒内，肽、蛋白质和寡核苷酸的许多功能取决于在皮秒至纳秒和更长时间内发生的结构波动。在每一种情况下，几乎没有实验技术可以解析演化系统的动态分子结构。我们的工作旨在通过开发能够收集振动光谱的超快拉曼光谱来纠正这一点，该振动光谱可以直接与生物分子中的光化学和热活化动力学的分子结构相关。该提案的具体目标#1将开发新的飞秒拉曼仪器，该仪器可以收集时间分辨率优于100 fs的高分辨率振动光谱。这种方法将被应用于获得新的理解的超快二聚化的嘧啶在DNA中的紫外光激发后，以及新的基本理解的量子力学性质的激发在核酸聚合物。在具体目标#2中，我们将开发新的方法来推动生物聚合物的热展开。这将允许众多的时间分辨光谱学工具，目前的应用仅限于光化学，解开皮秒到纳秒时间尺度上的热驱动二级结构变化的动力学。拉曼光谱学是唯一的定位，有助于这些领域，因为它的能力，收集振动光谱的生物聚合物在一个广泛的光谱窗口，而不干扰水环境。生物聚合物的拉曼光谱表现出特定的峰，这些峰是聚合物二级结构和侧链或核酸的特定环境的特征。因此，通过收集时间分辨的拉曼光谱的生化过程进行，我们可以确定的时间尺度的形成和动力学中间体的结构。这项实验工作将补充许多关于光激发DNA超快结构变化和更长时间尺度结构波动的理论预测。拟议的研究是重要的，因为光生物学的广度，可以解决这些技术和需要的实验探针的快速结构变化对生物学和人类健康的重要性。 公共卫生相关性（由申请人提供）：拟议的研究直接支持NIH的使命，帮助建立对紫外线损伤DNA的机制及其对公共卫生的影响的新认识。紫外线是地球上最普遍的环境诱变剂之一，对人类健康具有显著的有害影响。这些研究还将提高生物医学研究人员研究对蛋白质和DNA功能至关重要的快速结构运动的能力，从而加速新疗法的开发。