QUANTITATIVE STUDIES OF THE BIOTIN REGULATORY SYSTEM

原核生物和真核生物中基因表达的调控是 受到复杂的大分子组装体的影响。 这些功能 组装可以依赖于蛋白质-蛋白质，蛋白质-小分子， 和蛋白质-DNA相互作用。 生物素生物合成的调控 保留在E.大肠杆菌依赖于所有三种类型的 交互. 生物素系统的模型，基于 对大分子过程的定性理解， 有助于规范，已被提出。 生物素系统 提供了一个理想的模型系统， 个体大分子相互作用的生物学 复杂的基因调控系统 这项研究的目的是 建议是： 1.进行定量研究的相互作用， 生物素调节，以确定生物素的功能能量学。 生物素系统 2.调查负责的热力学驱动力 监管互动。 3.评价和修正生物素调节的定性模型 根据物理研究的结果， 测试系统的定量模型。 这些研究中使用的技术包括A。 定量DNA酶足迹技术测量DNA结合， B.平衡沉降测定亚基组装 性质，以及c.荧光光谱测量 异源蛋白-蛋白相互作用。 这些研究 生物素系统将有助于确定 单个生物素调节相互作用的物理化学， 整个系统的生物学，并可能揭示一般原则 与了解更复杂的基因调控相关的 系统. 相互作用的热力学特征， 此外，还能产生有关物理化学力的信息， 推动这些互动。

Regulation of gene expression in prokaryotes and eukaryotes is effected by complex macromolecular assemblies. Function of these assemblies can depend on protein-protein, protein-small molecule, and protein-DNA interactions. Regulation of biotin biosynthesis and retention in E. coli depends on all three types of interactions. A model for the biotin system, based on a qualitative understanding of the macromolecular processes that contribute to the regulation, has been proposed. The biotin system provides an ideal model system for relating the physical chemistry of individual macromolecular interactions to the biology of a complex gene regulatory system. The goals of this research proposal are to: 1. perform quantitative studies of the interactions that contribute to biotin regulation to define the functional energetics of the biotin system. 2. investigate the thermodynamic driving forces responsible for the regulatory interactions. 3. evaluate and revise the qualitative model for biotin regulation in light of the results of the physical studies and develop and test quantitative models for the system. The techniques that will be employed in these studies include a. the quantitative DNAse footprint technique to measure DNA binding, b. equilibrium sedimentation to determine of subunit assembly properties, and c. fluorescence spectroscopy to measure heterologous protein-protein interactions. These studies of the biotin system will help define the relationship between the physical chemistry of individual biotin regulatory interactions and the biology of the entire system and may reveal general principles that are relevant to understanding more complex gene regulatory systems. Thermodynamic characterization of the interactions will, moreover, yield information about the physical chemical forces that drive these interactions.