Centrifrugal analysis of complex hetero-interactions

复杂异质相互作用的离心分析

基本信息

批准号：
7014569
负责人：
JAMES L COLE
金额：
$ 17.53万
依托单位：
UNIVERSITY OF CONNECTICUT STORRS
依托单位国家：
美国
项目类别：
财政年份：
2004
资助国家：
美国
起止时间：
2004-03-01 至 2009-02-28
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7014569
关键词：
Internet automated data processing centrifugation computer data analysis computer program /software computer simulation computer system design /evaluation intermolecular interaction sedimentation equilibrium

项目摘要

DESCRIPTION (provided by applicant): Analytical ultracentrifugation is a rigorous method to characterize the stoichiometry, energetics and regulation of biomolecular interactions. Recent advances in instrument design and analysis methods have expanded the scope of sedimentation equilibrium measurements. However, this method has typically been applied to self-associating systems; studies of interactions between dissimilar partners (hetero-association) have been more limited by the complexity of experimental design, data analysis and the lack of appropriate software. Thus, many of the protein-protein interactions being discovered by high-throughput proteomics methods are not readily accessible for quantitative characterization by analytical ultracentrifugation. We propose to develop an integrated software package and new cell designs to facilitate studies of complex macromolecular interactions by sedimentation equilibrium. The package will simplify processing of primary data, fitting of experimental data to self- and hetero-association models by nonlinear least-squares algorithms and will integrate fitting results and graphical output. Analysis of hetero-associating systems is particularly challenging due to the complexity of the fitting models and contributions from multiple species present in solution. Our package will take advantage of the capabilities of the XL-I centrifuge by implementing global analysis of multiple data sets obtained at different sample concentrations, rotor speeds and detection methods (multiple absorption wavelengths and interference optics). In addition, the software will implement constraints on adjustable parameters as well as conservation of mass/signal algorithms to further confine fitted parameters to physically reasonable values. We also propose novel centrifuge cell designs to redistribute mass away from the cell boundaries so that errors in optical detection at the meniscus and base will be less significant in the conservation of mass calculations. The software will include modules for simulation of experimental data and error analysis routines to define the confidence intervals of the fitted parameters and permit visualization of the error surface. Our goal is to develop a robust, software package with a sophisticated user interface that is suitable for wide distribution to the scientific community. The analysis package, along with documentation and tutorials, will be freely available over the internet.

描述（由申请人提供）：分析超速离心是一种严格的方法，可以表征生物分子相互作用的化学计量，能量和调节。仪器设计和分析方法的最新进展扩大了沉积平衡测量的范围。但是，该方法通常应用于自我缔约系统。相互作用伙伴之间的相互作用（异质关联）之间的相互作用的研究受到了实验设计，数据分析和缺乏适当软件的复杂性的限制。因此，通过高通量蛋白质组学方法发现的许多蛋白质 - 蛋白质相互作用不容易通过分析性超速离心来定量表征。我们建议开发一个集成的软件包和新的细胞设计，以促进通过沉积平衡来研究复杂的大分子相互作用。该软件包将简化基本数据的处理，通过非线性最小二乘算法将实验数据拟合到自我和异核关系模型，并将整合拟合结果和图形输出。由于解决方案中存在的多个物种的拟合模型和贡献的复杂性，异质缔合系统的分析尤其具有挑战性。我们的软件包将通过对以不同样品浓度，转子速度和检测方法（多个吸收波长和干扰光学元件）获得的多个数据集进行全局分析来利用XL-I离心机的功能。此外，该软件将对可调参数以及质量/信号算法的保护实施约束，以将拟合参数限制在物理合理的值中。我们还提出了新型离心机细胞设计，以使质量从细胞边界重新分布，以便在弯弯；底部和基部的光学检测中的误差在守恒计算中的保存中不太重要。该软件将包括用于模拟实验数据和误差分析程序的模块，以定义拟合参数的置信区间并允许可视化误差表面。我们的目标是开发一个具有复杂用户界面的强大软件包，适用于科学界的广泛分发。分析软件包以及文档和教程将通过Internet免费获得。