TRANSIENT INFRARED PROBING OF PROTEIN FOLDNG AND CONFORMATIONAL DYNAMICS

蛋白质折叠和构象动力学的瞬态红外探测

• 批准号: 7598431
• 负责人: FENG GAI
• 金额: $ 3.97万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-09-01 至 2008-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7598431
• 关键词: Amino Acid Sequence; Area; Classification; Computer Retrieval of Information on Scientific Projects Database; Coupled; Detection; Development; Disease; Event; Fluorescence; Frequencies; Funding; Genes; Goals; Grant; Helix (Snails); Institution; Kinetics; Measures; Membrane Proteins; Modeling; Molecular; Molecular Conformation; Monitor; Motion; Numbers; Pathway interactions; Peptides; Play; Pliability; Protein Engineering; Proteins; Psychological Techniques; Range; Reaction Time; Research; Research Personnel; Resources; Role; Site; Source; Specificity; Spectroscopy, Fourier Transform Infrared; Spectrum Analysis; Structure; Structure-Activity Relationship; System; Time; Today; United States National Institutes of Health; Work; base; ear helix; infrared spectroscopy; insight; instrument; interest; millisecond; nanosecond; protein aggregation; protein folding; research study; structural biology; temperature jump; two-dimensional

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. The protein folding problem is considered to be one of the fundamental questions in structural biology. Folding research is today a very active area, where the experimental and theoretical techniques for probing folding at the molecular level are becoming more and more refined. The goal is to provide an experimental and theoretical basis for understanding and predicting protein folding pathways, the stable structures, and thermally and kinetically accessible conformation substates, given the primary amino acid sequence. A quantitative understanding of protein folding is apparently important for protein engineering. Understanding how proteins fold can also help to interpret quantitatively the structure-function relationships and folding related diseases. Furthermore, a predictive understanding of protein folding will accelerate the discovery of information contained in the large number of gene sequences that are now becoming available. It was proposed to develop instruments that are capable of triggering and probing conformational changes in proteins (and other molecular systems as well) on various timescales. Time-resolved infrared (IR) spectroscopy offers great flexibility and power for monitoring kinetic events on the molecular level with structure specificity and will be used to generate detailed structure interpretations of the transient species and their dynamics over the time range of interest. Using these instruments, we propose to study primarily how proteins fold. A detailed set of experiments are planned to gain detailed insight into the formation of protein secondary and tertiary structures. We are further extending current instruments and developing new instruments that are capable of triggering and probing conformational changes in proteins on various timescales. The specific aim of developing a nanosecond temperature-jump (T-jump) infrared spectrometer that can measure both transient kinetics at discrete frequencies and time-resolved spectra at discrete reaction times is being continued and extended. The microsecond FTIR coupled continuous-flow mixing and the millisecond FTIR coupled stopped-flow apparatus development is now available. The 2-dimensional (2D) correlation analysis has permitted site specific conformation studies and explorations of CN motions as a probe of dynamics. Studies of the helix-coil transition in alpha-helical peptides were performed, as well as studies of the stability and folding kinetics of beta-hairpin model peptides. We further work on the combination of the stop-flow apparatus with fluorescence detection and the incorporation of ATR spectroscopy into our IR capabilities to study membrane proteins. Another direction of this project will be the study of peptide/protein aggregation. Peptide and protein aggregation is the underlying cause of many diseases. This project is aimed to understand some fundamental aspects of peptide aggregation through a systematic approach. For example preliminary results on beta-hairpins suggest that the beta-turn plays a significant role in controlling the formation of beta-aggregates.

该副本是利用众多研究子项目之一 由NIH/NCRR资助的中心赠款提供的资源。子弹和 调查员（PI）可能已经从其他NIH来源获得了主要资金， 因此可以在其他清晰的条目中代表。列出的机构是 对于中心，这不一定是调查员的机构。 蛋白质折叠问题被认为是结构生物学中的基本问题之一。如今，折叠研究是一个非常活跃的领域，在该领域中，用于探测分子水平折叠的实验和理论技术正在变得越来越精致。目的是提供一个实验和理论基础，以理解和预测蛋白质折叠途径，稳定结构，以及在原发性氨基酸序列的情况下，在热和动力学上可访问的构象取代。对蛋白质折叠的定量理解对于蛋白质工程显然很重要。了解蛋白质如何折叠也可以帮助定量解释结构功能关系和相关疾病。此外，对蛋白质折叠的预测理解将加速发现现在已可用的大量基因序列中包含的信息。 提议开发能够触发和探测各个时间尺度上蛋白质（以及其他分子系统）的构象变化的仪器。时间分辨的红外（IR）光谱学为监测分子水平的动力学事件具有结构特异性提供了极大的灵活性和功能，并将用于在感兴趣的时间范围内生成瞬态物种及其动态的详细结构解释。使用这些仪器，我们建议主要研究蛋白质的折叠方式。计划进行详细的实验集，以详细了解蛋白质二级和三级结构的形成。 我们正在进一步扩展当前工具并开发能够触发和探测各个时间尺度蛋白质构象变化的新仪器。开发纳秒温度跳跃（T-JUMP）红外光谱仪的具体目的，该光谱仪可以在离散的频率下以离散的频率和时间分辨的光谱进行测量，并在离散反应时间延伸并扩展。现在可以使用微秒的FTIR耦合连续流混合和毫秒的FTIR耦合停止流动装置的开发。二维（2D）相关分析已允许位点特定的构象研究和CN运动的探索作为动力学探针。对α-螺旋肽中的螺旋线圈过渡进行了研究，并研究了β-毛发模型肽的稳定性和折叠动力学的研究。我们进一步致力于将荧光检测与荧光检测和ATR光谱合并到研究膜蛋白的能力中的结合。 该项目的另一个方向是研究肽/蛋白质聚集。肽和蛋白质聚集是许多疾病的根本原因。该项目的目的是通过系统的方法来了解肽聚集的一些基本方面。例如，对β发pin的初步结果表明，β-转弯在控制β-聚集物的形成中起着重要作用。