Thermodynamic Analysis of Transmembrane Beta Barrel Self-Association

跨膜 Beta 桶自缔合的热力学分析

• 批准号: 0423807
• 负责人: Karen Fleming
• 金额: --
• 依托单位: Johns Hopkins University
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-08-01 至 2009-07-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=0423807&HistoricalAwards=false
• 关键词: Thermodynamic; Analysis; Transmembrane; Beta; Barrel

The goals of this research are to elucidate an understanding of the molecular principles of membrane protein interactions. These interactions are important for cellular function, but they are currently poorly understood. To accomplish this goal, the stability of the transmembrane beta-barrel protein, OMPLA, will be determined. The influence of calcium and substrate, two biological effector molecules, on OMPLA self-association will also be quantified. To meet these goals, two sets of experimental studies will be carried out. Sedimentation equilibrium analytical ultracentrifugation will be used to measure OMPLA self-association equilibrium constants in the presence and absence of substrate and calcium. Spectroscopic techniques will be used to determine the strength of calcium binding to the monomeric and dimeric forms of OMPLA. To understand the mechanisms by which these three effects regulate the OMPLA activity, the thermodynamic linkage relationships between them will be determined. This thermodynamic data will be used to construct appropriate mathematical equations that describe the species populations, from which one can gain a greater understanding of the OMPLA species that are biologically relevant. The structure/energy relationships for membrane protein interactions have only been extensively studied in two helical membrane proteins. These results will be the first for a transmembrane beta-barrel protein and will facilitate a comparison of interaction motifs and principles between transmembrane alpha-helical and beta-barrel proteins. This project will allow students to gain expertise in analytical ultracentrifugation as well as mathematics. To train young scientists, undergraduate students will be given the opportunity for a meaningful research experience. At the same time, this will have the dual benefit of offering graduate students the opportunity to learn how to mentor students and troubleshoot projects.

这项研究的目的是阐明膜蛋白相互作用的分子原理。这些相互作用对细胞功能很重要，但目前人们对它们知之甚少。为了实现这一目标，将测定跨膜β桶蛋白OMPLa的稳定性。钙和底物这两个生物效应分子对OMPLA自结合的影响也将被量化。为了达到这些目标，将进行两套实验研究。沉降平衡分析超速离心法将被用来在底物和钙的存在和不存在的情况下测量OMPLA的自缔合平衡常数。光谱技术将被用来确定钙与OMPLA单体和二聚体的结合强度。为了了解这三种效应调节OMPLA活性的机制，我们将确定它们之间的热力学联系。这些热力学数据将被用来构建描述物种种群的适当数学方程，从中人们可以更好地了解与生物相关的OMPLA物种。膜蛋白相互作用的结构/能量关系仅在两种螺旋膜蛋白中得到了广泛的研究。这些结果将是跨膜β-桶蛋白的第一次，并将有助于比较跨膜α-螺旋和β-桶蛋白之间的相互作用基序和原理。这个项目将使学生获得分析超速离心学和数学方面的专业知识。为了培养年轻科学家，本科生将有机会获得有意义的研究经历。与此同时，这将有双重好处，即为研究生提供学习如何指导学生和解决项目问题的机会。