Development of Biosensor Technology for Protein Interactions

蛋白质相互作用生物传感器技术的发展

• 批准号: 7967910
• 负责人: PETER SCHUCK
• 金额: $ 3.05万
• 依托单位: NATIONAL INSTITUTE OF BIOMEDICAL IMAGING AND BIOENGINEERING
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/7967910
• 关键词: Accounting; Affinity; Binding; Binding Sites; Biological Assay; Biosensor; Chemicals; Computer software; Data; Data Set; Development; Experimental Models; Goals; Heterogeneity; Immobilization; Kinetics; Knowledge; Membrane Proteins; Microscopic; Modeling; Noise; Polymers; Population; Property; Proteins; Role; Scientist; Site; Solutions; Surface; Surface Plasmon Resonance; System; Technology; Time; chemical property; crosslink; data acquisition; macromolecule; new technology; tool

Due to surface rugosity, the physical and chemical microenvironment at the surface is heterogeneous. Therefore, proteins that have a single class of binding sites in solution can show a dispersion in the binding properties once chemically crosslinked to the surface. As a tool to study this ensemble of surface sites, we have previously introduced a computational approach to determine distributions of affinity and kinetic binding site parameters from experimental surface binding data. This allowed us, for the first time, to account simultaneously for the two most commonly encountered experimental problems of surface binding when using biosensors for characterizing protein interactions, site heterogeneity and mass transport limitation, and thereby model experimental data consistently to within the level of noise of data acquisition. This fully exploits the high sensitivity of surface plasmon resonance biosensors for the study of protein interactions, and provides information of the surface binding sites with unprecedented level of detail. Further applications of this approach to more protein interactions have solidified our knowledge of surface site heterogeneity. We have developed a stand-alone software that allows other scientist to import experimental surface binding kinetics data sets and compute the functional distribution of sites. We have confirmed with a model protein system how the direct protein immobilization can cause heterogeneity and sub-populations with different affinity. We have implemented software for routine analysis with a solution competition assay that quantifies protein interactions in solution.

由于表面皱纹，表面的物理和化学微环境是异质的。 因此，在溶液中具有单一类结合位点的蛋白质可以在化学上与表面进行化学交联的结合特性中表现出分散体。 作为研究表面位点整体的工具，我们先前曾引入一种计算方法，以确定从实验表面结合数据中确定亲和力和动力学结合位点参数的分布。 这使我们第一次可以同时考虑两个最常见的最常见的表面结合实验问题，当时使用生物传感器来表征蛋白质相互作用，现场异质性和质量传输限制，从而将实验数据始终如一地模拟到数据采集噪声范围内。 这充分利用了表面等离子体共振生物传感器对蛋白质相互作用的高灵敏度，并提供了前所未有的细节水平的表面结合位点的信息。 这种方法在更多蛋白质相互作用中的进一步应用巩固了我们对表面位点异质性的了解。 我们已经开发了一个独立的软件，该软件允许其他科学家进口实验表面结合动力学数据集并计算位点的功能分布。 我们已经通过模型蛋白系统证实了直接蛋白固定化如何导致具有不同亲和力的异质性和亚群。 我们已经通过解决方案中的蛋白质相互作用的解决方案竞争测定法实施了用于例行分析的软件。