CORE--MICROANALYSIS OF PEPTIDES AND PROTEINS

核心--肽和蛋白质的微量分析

• 批准号: 6098644
• 负责人: Susan M Lunte
• 金额: $ 13.26万
• 依托单位: UNIVERSITY OF KANSAS LAWRENCE
• 依托单位国家: 美国
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-08-01 至 1999-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6098644
• 关键词: analytical chemistry; analytical method; binding proteins; biomedical facility; calcium transporting ATPase; calmodulin; capillary electrophoresis; electrospray ionization mass spectrometry; fluorescence spectrometry; gel electrophoresis; glutamates; high performance liquid chromatography; mass spectrometry; membrane transport proteins; oxidation; posttranslational modifications; protein purification; protein sequence; protein structure function

There is a considerable amount of evidence that oxidation of proteins by reactive oxygen species occurs during aging. One of the major objectives of this Program Project is to correlate chemical changes shown to occur in vitro with isolated proteins with oxidative modifications found in aged tissue. Toward this end, the role of several different ROS in the oxidation of CaM, SR and plasma Ca-ATPase, Na+/Ca2+ exchanger and the glutamate binding proteins will be investigated. Although these five proteins differ considerably in their physical properties and biological function, the basic analytical needs to detect the oxidative modifications are the same. Therefore, it is proposed that a Core Facility for the microanalysis of peptides and proteins be set up within the Center for Bioanalytical Research (CBAR) at the University of Kansas. The faculty members of CBAR already have considerable experience in the trace determination of peptides and amino acids, and the existence of this facility within CBAR will provide project members with access to this expertise. The rationale for the Core Facility is based on the following: (1) cost savings by making the instrumentation accessible to all investigators, eliminating redundancies in equipment in individual investigator's laboratories, (2) scientists employed by the Core Facility will spend their time exclusively on the development of analytical methodology for the Program Project, allowing the Program Project members to concentrate on the biological aspects of aging, (3) access to additional instrumentation and expertise within the Center for Bioanalytical Research (4) because of the centralized nature of the facility, it will be easier to incorporate new developments in microanalytical methodology into the Program Project. Because of the potentially small amounts of protein involved in this project, a major emphasis of the facility will be the use of microcolumn-based separation techniques. These include microbore and capillary LC and capillary electrophoresis. Whenever possible, mass spectrometry will be employed as the method of structural identification. However, fluorescence and/or electrochemical detection methods will be employed for identification of specific modifications when appropriate.

有大量证据表明，蛋白质氧化 活性氧在衰老期间发生。 主要目标之一 这个程序项目的是将显示发生的化学变化相关联 体外与分离的蛋白质具有氧化修饰 组织。 为此，几种不同的ROS在 CAM，SR和血浆CA-ATPase，Na+/Ca2+交换器的氧化和 将研究谷氨酸结合蛋白。 虽然这五个 蛋白质的物理特性和生物学差异很大 功能，基本分析需要检测氧化修饰 是一样的。 因此，有人提出了一个核心设施 在中心内建立肽和蛋白质的微分析 堪萨斯大学的生物分析研究（CBAR）。 教师 CBAR成员在痕迹方面已经有丰富的经验 测定肽和氨基酸，并存在 CBAR内的设施将为项目成员提供访问权限 专业知识。 核心设施的理由是基于以下基础： （1）通过使所有人都可以访问仪器来节省成本 调查人员，消除了单个设备的冗余 研究人员的实验室（2）核心设施雇用的科学家 将花时间专门用于分析的发展 计划项目的方法学，允许计划项目成员 要专注于衰老的生物学方面，（3） 中心内的其他仪器和专业知识 生物分析研究（4），因为 设施，将新的发展纳入 微分析方法进入程序项目。 因为 该项目可能涉及的少量蛋白质，这是一个主要的 该设施的重点将是使用基于微柱的分离 技术。 其中包括微孔和毛细管LC和毛细管 电泳。 只要有可能，质谱将被用作 结构鉴定的方法。 但是，荧光和/或 电化学检测方法将用于识别 适当时进行特定修改。