Enthalpy arrays for thermodynamic characterization

用于热力学表征的焓阵列

• 批准号: 7628512
• 负责人: Richard H. Bruce
• 金额: $ 36.63万
• 依托单位: PALO ALTO RESEARCH CENTER
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-06-01 至 2011-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7628512
• 关键词: Address; Affinity; Area; Basic Science; Binding; Biochemical; Biochemistry; Biological Assay; Biology; CD209 gene; Calorimetry; Carbohydrates; Cells; Chemicals; Collaborations; Commit; Consumption; DNA; Data; Detection; Development; Discipline; Disclosure; Engineering; Ensure; Entropy; Free Energy; Funding; Generic Drugs; Goals; HIV; HIV Infections; HIV Protease; HIV Protease Inhibitors; Immobilization; Immune system; Label; Letters; Life Cycle Stages; Ligand Binding; Ligands; Measurement; Measures; Mechanics; Methods; Microfabrication; Modification; Molecular; Noise; Nucleic Acids; Physics; Production; Proteins; Proteomics; RNA; Reaction; Reagent; Research; Research Personnel; Sampling; Screening procedure; Senior Scientist; Surface; System; Techniques; Technology; Thermodynamics; Time; Titrations; Work; assay development; base; chemical reaction; cost; design; detector; drug discovery; enthalpy; evaporation; experience; improved; inhibitor/antagonist; instrument; interest; intermolecular interaction; meetings; microcalorimetry; multidisciplinary; neutralizing antibody; programs; receptor; sensor; small molecule; stoichiometry; technology development; tool

DESCRIPTION (provided by applicant): A generic method to characterize molecular interactions with no assay development would substantially benefit both functional proteomics research and drug discovery; however, none of the available approaches offers the efficient detection and low reagent consumption needed for the large number of measurements. This proposal addresses developing a sensitive enthalpic assay tool to detect bimolecular interactions involving proteins, ligands, RNA, DNA and other molecules of interest without requiring any special assay development. In particular, the assay does not require development of molecule-specific inhibitors, probes, detectors, attachment of labels, immobilization on surfaces, or chemical modifications of the molecules. The assay is based on direct detection of the enthalpy released during interaction of the species of interest. The detector, a nanocalorimeter, will enable measurement of interactions for molecular concentrations in the low micro-molar range using sample volumes of 200 nl. The technical goal of this work is to develop a detector that can detect enthalpies in the nanocalorie range with an effective measurement time of six seconds. This represents nearly three orders of magnitude reduction in the required sample and measurement time compared with conventional microcalorimetry. We propose to develop an array of 96 detectors and use it to study key aspects of HIV infection and life cycle. In addition to detecting interactions the enthalpy array can be used to perform titrations to determine binding constants. The effort integrates several engineering disciplines with applied physics and biochemistry. We have developed the initial proof of concept through a multidisciplinary effort bridging microfabrication with several engineering disciplines (mechanical, electrical chemical and system) as well as applied physics. We are collaborating with several senior biochemists to ensure that the system meets their scientific needs.

描述（由申请人提供）：无需开发测定即可表征分子相互作用的通用方法将大大有利于功能蛋白质组学研究和药物发现；然而，没有一种可用的方法能够提供大量测量所需的高效检测和低试剂消耗。该提案旨在开发一种灵敏的焓测定工具来检测涉及蛋白质、配体、RNA、DNA 和其他感兴趣分子的双分子相互作用，而无需任何特殊的测定开发。特别是，该测定不需要开发分子特异性抑制剂、探针、检测器、标记的附着、表面固定或分子的化学修饰。该测定基于直接检测感兴趣物种相互作用期间释放的焓。该探测器是一种纳量热计，能够使用 200 nl 的样品体积测量低微摩尔范围内分子浓度的相互作用。这项工作的技术目标是开发一种能够检测纳卡范围内的焓的探测器，有效测量时间为六秒。与传统的微量热法相比，这意味着所需的样品和测量时间减少了近三个数量级。我们建议开发一系列 96 个探测器，并用它来研究 HIV 感染和生命周期的关键方面。除了检测相互作用之外，焓阵列还可用于执行滴定以确定结合常数。这项工作将多个工程学科与应用物理和生物化学相结合。我们通过多学科努力，将微加工与多个工程学科（机械、电化学和系统）以及应用物理学结合起来，开发了初步的概念验证。我们正在与几位资深生物化学家合作，以确保该系统满足他们的科学需求。

项目成果

Higher throughput calorimetry: opportunities, approaches and challenges.

• DOI: 10.1016/j.sbi.2010.09.001
• 发表时间: 2010-10
• 期刊: CURRENT OPINION IN STRUCTURAL BIOLOGY
• 影响因子: 6.8
• 作者: Torres, Francisco E.;Recht, Michael I.;Coyle, Joseph E.;Bruce, Richard H.;Williams, Glyn
• 通讯作者: Williams, Glyn

Rapid mixing of sub-microlitre drops by magnetic micro-stirring.

• DOI: 10.1039/c1lc20354a
• 发表时间: 2011-10-07
• 期刊: Lab on a chip
• 影响因子: 6.1
• 作者: De Bruyker D;Recht MI;Bhagat AA;Torres FE;Bell AG;Bruce RH
• 通讯作者: Bruce RH