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.

项目概要 测量分子与细胞膜蛋白的相互作用并量化相互作用动力学 实时对于理解许多细胞过程、验证生物标志物和筛选至关重要 药物。这是因为膜蛋白负责细胞的许多重要的细胞功能， 包括与其他细胞的通信、感知周围环境以及运输分子 以及进出细胞的离子。它们还包含近 60% 的当前药物靶点。然而，开发这样的 能力一直是一个艰巨的挑战，特别是对于小分子来说，因为大多数传统的结合 动力学测量技术基于分子质量的检测，分子质量随着分子质量的增加而减小 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。 为了解决未满足的需求，该项目将开发机械放大光学检测技术。 该技术基于基本热力学原理，即细胞中的机械变形 当细胞上发生分子结合事件时，膜就会形成。通过准确监控 机械变形，因此可以确定大分子和小分子与 细胞上的膜蛋白。这种新策略为小结合信号提供机械放大， 结合新颖的成像技术和信号处理算法来跟踪细胞变形 亚纳米精度，使得检测分子与膜蛋白的相互作用成为可能。它还 允许通过分析不同细胞之间的结合动力学变异性来研究细胞的异质性 细胞。最后，该技术可以研究难以或不可能分离的膜蛋白 来自具有完整天然结构和活性的细胞。 该项目的具体目标是1）建立数据采集和分析算法以准确地 检测分子结合诱导的细胞膜变形，2）开发单细胞的高通量系统 细胞结合动力学分析，3）开发用于研究低密度膜的低噪声成像技术 蛋白质，4) 验证新技术的性能和可用性。该项目的成功将 为生物医学研究界和工业界提供一种研究基本细胞过程的新工具， 验证生物标志物以及筛选新药。