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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

基本信息

批准号：
8112687
负责人：
David W McCamant
金额：
$ 18.45万
依托单位：
UNIVERSITY OF ROCHESTER
依托单位国家：
美国
项目类别：
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/8112687
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 Pyrimidines Raman Spectrum Analysis Research Research Design Research Methodology 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

项目摘要

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在飞秒（10-15秒）到纳秒（10-9秒）时间尺度上发生在生物系统中的结构变化。紫外线对核酸的光损伤发生在100飞秒内，多肽、蛋白质和寡核苷酸的许多功能依赖于皮秒到纳秒甚至更长时间内发生的结构波动。在每种情况下，很少有实验技术可以解决进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与生物分子光化学和热激活动力学的分子结构直接相关的振动光谱的超快拉曼光谱来纠正这一问题。本提案的具体目标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。该方法将应用于对紫外激发后DNA中嘧啶的超快二聚化的新认识，以及对核酸聚合物中激发的量子力学性质的新基本认识。具体目标#2，我们将开发新的方法，以脉冲启动热展开的生物聚合物。这将允许时间分辨光谱的许多工具，目前的应用仅限于光化学，揭示在皮秒到纳秒时间尺度上热驱动的二级结构变化的动力学。拉曼光谱在这些领域具有独特的地位，因为它能够在宽光谱窗口内收集生物聚合物的振动光谱，而不会受到水环境的干扰。生物聚合物的拉曼光谱表现出特定的峰，这些峰是聚合物的二级结构和侧链或核酸的特定环境的特征。因此，通过收集生化过程进行时的时间分辨拉曼光谱，我们可以确定形成的时间尺度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的超快结构变化和更长的时间尺度结构波动的理论预测。这项研究具有重要意义，因为光生物学的广度可以通过这些技术来解决，并且需要对生物学和人类健康重要的快速结构变化进行实验探测。