Measuring small molecule interactions with membrane proteins on single cells via detecting nanometer scale membrane deformations

通过检测纳米级膜变形来测量小分子与单细胞膜蛋白的相互作用

• 批准号: 9365667
• 负责人: NONGJIAN TAO
• 金额: $ 30.39万
• 依托单位: ARIZONA STATE UNIVERSITY-TEMPE CAMPUS
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-08-01 至 2021-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9365667
• 关键词: Accounting; Address; Affinity; Algorithmic Analysis; Algorithms; Binding; Binding Proteins; Biological Markers; Biomedical Research; Cell membrane; Cell physiology; Cell surface; Cells; Communication; Communities; Data Analyses; Dependence; Detection; Drug Targeting; Environment; Evaluation; Event; Fourier Transform; Goals; Image; Imaging technology; Immobilization; Individual; Industry; Ions; Kinetics; Label; Liquid substance; Measurement; Measures; Mechanics; Membrane; Membrane Proteins; Methods; Microscopic; Molecular; Monitor; Nature; Noise; Optics; Performance; Pharmaceutical Preparations; Preclinical Drug Evaluation; Preparation; Process; Protein Conformation; Proteins; Protocols documentation; Sampling; Signal Transduction; Specificity; Structure; Surface; System; Technology; Thermodynamics; Time; Validation; Work; base; cell fixation; contrast imaging; data acquisition; density; drug discovery; experimental study; high throughput analysis; molecular mass; molecular scale; nanometer; nanoscale; new technology; novel; novel therapeutics; optical imaging; protein function; risk minimization; sample fixation; screening; signal processing; small molecule; success; temporal measurement; tool; usability

PROJECT SUMMARY Measuring interactions of molecules with membrane proteins on cells and quantifying the interaction kinetics in real time are critical for understanding many cellular processes, for validating biomarkers, and for screening drugs. This is because membrane proteins are responsible for many important cellular functions of cells, including communication with other cells, sensing the surrounding environment, and transporting molecules and ions in and out of cells. They also comprise nearly 60% of current drug targets. However, developing such a capability has been a difficult challenge, especially for small molecules, because most traditional binding kinetics measurement technologies are based on the detection of molecular mass, which diminishes with the size of the molecule. Small molecules are the most important forms of drugs, accounting for over 70% of all the drugs developed to date. To address the unmet need, this project will develop a mechanically amplified optical detection technology. The technology is based on a basic thermodynamics principle that a mechanical deformation in the cell membrane occurs when a molecular binding event takes place on the cell. By accurately monitoring the mechanical deformation, one can thus determine the kinetics of both large and small molecule binding with membrane proteins on cells. This new strategy provides mechanical amplification to small binding signals, which, together with a novel imaging technology and signal processing algorithm to track cell deformation with sub-nanometer accuracy, make it possible to detect molecular interactions with membrane proteins. It also allows the study of heterogeneous nature of cells by analyzing the binding kinetics variability between different cells. Finally, the technology allows the study of membrane proteins that are difficult or impossible to isolate from cells with intact native structures and activities. The specific aims of the project are to 1) establish data acquisition and analysis algorithms to accurately detect molecular binding-induced cell membrane deformation, 2) develop a high-throughput system for single cell binding kinetics analysis, 3) develop a low-noise imaging technology for studying low-density membrane proteins, and 4) validate the performance and usability of the new technology. The success of the project will lead to a new tool for biomedical research community and industry for studying basic cellular processes, for validating biomarkers, and for screening new drugs.

项目摘要 测量分子与膜蛋白在细胞上的相互作用，并量化相互作用动力学 实时对于理解许多蜂窝过程，验证生物标志物和筛查至关重要 毒品。这是因为膜蛋白负责细胞的许多重要细胞功能， 包括与其他单元的通信，感测周围环境和运输分子 和离子进出细胞。它们还占当前药物靶标的近60％。但是，开发这种情况 功能一直是一个困难的挑战，尤其是对于小分子，因为大多数传统的绑定 动力学测量技术基于检测分子质量，该质量会减少 分子的大小。小分子是最重要的药物形式，占所有药物的70％以上 迄今为止开发的药物。 为了满足未满足的需求，该项目将开发机械放大的光学检测技术。 该技术基于基本的热力学原理，该原理是细胞中的机械变形 当分子结合事件发生在细胞上时，会发生膜。通过准确监视 因此，机械变形，因此可以确定大和小分子结合的动力学 细胞上的膜蛋白。这种新策略为小型绑定信号提供了机械扩增， 它以及一种新型的成像技术和信号处理算法，以跟踪细胞变形 亚纳米的精度，可以检测与膜蛋白的分子相互作用。也是如此 可以通过分析不同的动力学变异性来研究细胞的异质性质 细胞。最后，该技术允许研究难以或不可能分离的膜蛋白 来自具有完整的天然结构和活性的细胞。 该项目的具体目的是1）建立数据采集和分析算法以准确 检测分子结合诱导的细胞膜变形，2）为单个开发高通量系统 细胞结合动力学分析，3）开发一种用于研究低密度膜的低噪声成像技术 蛋白质以及4）验证新技术的性能和可用性。该项目的成功将 导致生物医​​学研究社区和行业的新工具，用于研究基本的细胞过程 验证生物标志物，用于筛查新药。