SPECTROPOLARIMETER: PROTEIN: STRUCTURES & FOLDING MECHANISM

该子项目是利用由NIH/NCRR资助的共享仪器赠款提供的资源的许多研究子项目之一。子项目和研究者（PI）可能从另一个NIH来源获得主要资金，因此可以在其他CRISP条目中表示。列出的机构是用于资助的，不一定是研究者的机构。描述（由申请人提供）：三名NCSU科学家在蛋白质-蛋白质或蛋白质-核酸相互作用是主要组成部分的领域工作，建议购买能够进行稳态和停流圆二色性（CD）研究的分光偏振仪。该仪器由Applied Photophysics制造，旨在取代和升级1989年购买的现有和过时的Jasco CD。现有的仪器仍然是NCSU校园内唯一的CD仪器，并被校园内的几个教师大量使用。然而，为了获取足够的数据，将仪器保持在工作状态变得越来越困难。现有的仪器在200 nm以下不能提供可靠的数据，温度控制差，软件和计算机陈旧，没有停流或滴定功能。在过去的三年里，该仪器需要数千美元的维修费用才能保持功能。拟议的仪器将使蛋白质和核酸构象的研究，并提供有关蛋白质和/或核酸的稳定性以及蛋白质折叠机制的信息。结合曲线的测量将允许确定结合常数和化学计量，将获得对圆二色性参数的热依赖性，并将测量动力学反应。该仪器将促进研究人员目前的实验方法，其中包括NMR光谱学，荧光光谱学，X射线晶体学，差示扫描和等温滴定量热法，圆二色性，质谱法，以及其他生物化学和生物物理技术。稳态和停流分光光度计将补充NIH资助的结构和功能研究，重点是了解半胱天冬酶的折叠和组装，参与细菌反应和保护的过渡态调节蛋白，以及修饰的核苷对RNA和RNA的表征：肽和RNA：金属相互作用。

This subproject is one of many research subprojects utilizing the resources provided by a Shared Instrumentation 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 grant, which is not necessarily the institution for the investigator. DESCRIPTION (provided by applicant): Three NCSU scientists working in fields in which protein-protein or protein-nucleic acids interactions are a primary component, propose to purchase a spectropolarimeter capable of steady-state and stopped-flow circular dichroism (CD) studies. The instrument, manufactured by Applied Photophysics, is intended as a replacement and upgrade for an existing and outdated Jasco CD purchased in 1989. The existing instrument remains the only CD instrument on the NCSU campus and is used heavily by several faculty across the campus. However, it becomes increasingly difficult to maintain the instrument in working condition in order to acquire adequate data. The existing instrument will not provide reliable data below 200 nm, the temperature control is poor, the software and computer are antiquated, and it does not have stopped-flow or titration capabilities. In the past three years, the instrument has required several thousand dollars in repair costs to remain functional. The proposed instrument will enable the study of protein and nucleic acid conformations and provide information on protein and/or nucleic acid stability as well as protein folding mechanisms. Measurements of binding profiles will allow the determination of binding constants and stoichiometries, thermal dependence to circular dichroism parameters will be obtained, and kinetic reactions will be measured. The instrument will facilitate the current experimental approaches of the investigators which include NMR spectroscopy, fluorescence spectroscopy, X-ray crystallography, differential scanning and isothermal titration calorimetry, circular dichroism, mass spectrometry, and other biochemical and biophysical techniques. The steady-state and stopped-flow spectropolarimeter will complement NIH funded structural and functional studies focused on understanding the folding and assembly of caspases, transition state regulator proteins involved in bacterial response and protection, and characterization of modified nucleoside contributions to RNA and to RNA:peptide and RNA:metal interactions.