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HTS of small molecule-protein interactions (RMI)

小分子-蛋白质相互作用 (RMI) 的 HTS

基本信息

批准号：
7477886
负责人：
DALE NORMAN LARSON
金额：
$ 66.04万
依托单位：
CHARLES STARK DRAPER LABORATORY
依托单位国家：
美国
项目类别：
财政年份：
2005
资助国家：
美国
起止时间：
2005-09-23 至 2010-07-31
项目状态：
已结题

项目摘要

DESCRIPTION (provided by applicant): Understanding the selectivity of small molecule binding is a central question in drug development. It impinges on both determining which desirable targets might respond to the compound as well as which unintended targets might lead to toxicity. Currently these are difficult questions to answer because they require the evaluation of the small molecule's interactions with many proteins, which in turn requires methods for testing many proteins and for detecting and characterizing the interaction. Protein microarrays offer a compelling method for presenting many proteins for testing, but the methods currently available for assessing small molecule binding require modifying the small molecule with a label. This labeling frequently results in changes to the small molecule's binding performance and can be difficult. A technology that enables researchers to address these questions in a high-throughput manner will accelerate the transition of discoveries into the clinic. In this project, we propose to bring two technologies together that will result in a high-throughput system to detect and characterize small molecule-protein interactions using a label-free system that is extremely sensitive, quantitative and provides information on binding kinetics. The two technologies are a label-free multiplexed detection system that is based on nanohole arrays and a protein microarray system, nucleic acid programmable protein array (NAPPA). The nanohole array sensor, technology is a detection method based on a novel photonics device whose key characteristics are: single molecule sensitivity, a very high level of multiplexing, label-free detection, and the generation of kinetic binding data. The NAPPA technology is a method for the miniaturized presentation of thousands of proteins based on in situ expression of protein from cDNAs printed onto a surface. NAPPA addresses the key constraints associated with other protein microarray systems, namely, protein stability and storage, elimination of the need for protein purification, and captures 1000 fold more protein than other approaches. This project also makes use of a library of expression ready cDNA clones for human proteins that is being assembled by the Harvard Institute of Proteomics. These technologies have the potential to produce a valuable tool in lead identification and optimization.

描述（由申请人提供）：了解小分子结合的选择性是药物开发中的一个中心问题。它影响到确定哪些理想的目标可能对化合物产生反应，以及哪些非预期的目标可能导致毒性。目前，这些问题很难回答，因为它们需要评估小分子与许多蛋白质的相互作用，这反过来又需要用于测试许多蛋白质以及用于检测和表征相互作用的方法。蛋白质微阵列提供了一种引人注目的方法来呈现许多蛋白质进行测试，但目前可用于评估小分子结合的方法需要用标签修饰小分子。这种标记经常导致小分子结合性能的变化，并且可能是困难的。一项使研究人员能够以高通量方式解决这些问题的技术将加速发现向临床的转变。在这个项目中，我们建议将两种技术结合在一起，这将导致一个高通量的系统，使用一个无标记的系统，是非常敏感的，定量的，并提供有关结合动力学的信息来检测和表征小分子-蛋白质相互作用。这两种技术是基于纳米孔阵列的无标记多重检测系统和蛋白质微阵列系统，核酸可编程蛋白质阵列（NAPPA）。纳米孔阵列传感器技术是一种基于新型光子器件的检测方法，其主要特征是：单分子灵敏度，非常高的多路复用水平，无标记检测和动力学结合数据的生成。NAPPA技术是一种基于从印刷到表面上的cDNA原位表达蛋白质来微型化呈现数千种蛋白质的方法。NAPPA解决了与其他蛋白质微阵列系统相关的关键限制，即蛋白质稳定性和储存，消除了对蛋白质纯化的需要，并捕获了比其他方法多1000倍的蛋白质。该项目还利用了一个由哈佛蛋白质组学研究所正在组装的人类蛋白质表达准备cDNA克隆库。这些技术有可能产生一个有价值的工具，在铅识别和优化.