Development of Cavity-Enhanced Single-Molecule Electronic and Vibrational Spectroscopy for Mechanistic Studies of Biomolecules

用于生物分子机理研究的腔增强单分子电子和振动光谱学的发展

• 批准号: 10251006
• 负责人: Randall H Goldsmith
• 金额: $ 28.04万
• 依托单位: UNIVERSITY OF WISCONSIN-MADISON
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-09-05 至 2024-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10251006
• 关键词: Biochemical; Biological; Chemicals; Cobalt; Complex; Coupling; Development; Devices; Diagnostic; Dyes; Environment; Evaluation; Fiber; Film; Fluorescence; Geometry; Glass; In Vitro; Individual; Investigation; Label; Learning; Light; Measurement; Metals; Methods; Microbubbles; Molecular; Monitor; Optics; Performance; Polymers; Positioning Attribute; Production; Property; Pump; Raman Spectrum Analysis; Sampling; Series; Signal Transduction; Specificity; Spectrum Analysis; Surface; System; Techniques; Technology; Temperature; Testing; Thermometers; Time; Vitamin B 12; absorption; aqueous; biomaterial compatibility; biophysical techniques; chemical bond; chemical reaction; design; indexing; insight; instrumentation; metalloenzyme; molecular dynamics; nanoparticle; oxidation; particle; photonics; plasmonics; silicon nitride; single molecule; tool; vibration

Development of Cavity-Enhanced Single-Molecule Electronic and Vibrational Spectroscopy for Mechanistic Studies of Biomolecules Single-molecule (SM) measurements are a powerful mechanistic tool because they allow multi-step unsynchro- nized dynamics to be directly observed. However, most SM observations rely on fluorescence, which lacks the sensitivity to determine oxidation state, the chemical specificity to elucidate distortion of a particular chemical bond, and requires a fluorescent label. Such information would revolutionize how biochemical mechanisms are determined and could be provided by a method of performing electronic absorption and vibrational spectroscopy on single operational biomolecules. However, surface-enhanced Raman spectroscopy (SERS) is not is suited for probing complex biomolecules, as the method requires intimate contact between the part of the biomolecule to be probed (which may be at the interior), and a metal surface. Similarly, methods exist for performing SM elec- tronic absorption spectroscopy but they lack the required sensitivity or biocompatibility for biomolecules. Thus, a new method is needed to allow SM investigations of in vitro molecular dynamics for mechanistic investigations. We propose the use of optical microcavities as platforms for ultrasensitive SM electronic and vibrational spectroscopy. In one geometry, microcavities are used as highly sensitive thermometers, capable of detecting the heat dissipated by non-fluorescent molecules upon photoexcitation. In this way, non-fluorescent and potentially even weakly absorbing spectral features, such as those diagnostic of the coordination environment of a metal- loenzyme can be elucidated. In a second complimentary geometry we take advantage of the Purcell Effect, which can significantly enhance scattering rates in optical microcavities with small mode volumes and high Quality factors. While SERS requires essentially Van der Waals contact with a plasmonic surface, the microcavity en- hancement can act at a distance of up to ~100 nm from a dielectric surface, making it suitable for probing bio- molecules without significant perturbation. We have now demonstrated the core concepts behind these two strat- egies. In Specific Aims 1-3, we will bring online and evaluate three new microcavity systems that promise to significantly enhance our measurement capacity enough to lay a concrete path to biomedical applications: planar silicon nitride ring resonators (SA 1), fiber Fabry-Perot microcavities (SA 2), and silicon nitride nanobeams (SA3). In all cases we will perform spectroscopy on a series of particles and molecules of increasing challenge, pushing toward the monitoring of a single working metalloenzyme. Supporting calculations suggest that these new resonator geometries will increase our molecular signals by orders of magnitude. Our long-term objective is to bring a new, highly informative, and even disruptive biophysical technique to bear on biological molecules to understand how they operate, change in time, are regulated, and fail.

光腔增强单分子电子和振动光谱研究进展 生物分子的机理研究 单分子（SM）测量是一种强大的机械工具，因为它们允许多步不同步测量。 可以直接观察到的动态。然而，大多数SM观测依赖于荧光，而荧光缺乏 确定氧化态的灵敏度，阐明特定化学品畸变的化学特异性 键，并且需要荧光标记。这些信息将彻底改变生物化学机制 可以通过进行电子吸收和振动光谱的方法来确定和提供 在单一的生物分子上。然而，表面增强拉曼光谱（Sers）不适合于 探测复杂的生物分子，因为该方法需要生物分子的一部分之间的紧密接触， 被探测（其可以在内部）和金属表面。类似地，存在用于执行SM同步的方法。 电子吸收光谱，但它们缺乏所需的灵敏度或生物分子的生物相容性。因此，在本发明中， 需要一种新的方法来允许SM研究用于机理研究的体外分子动力学。 我们建议使用光学微腔作为超灵敏SM电子和振动的平台， 谱在一种几何形状中，微腔用作高灵敏度温度计，能够检测温度。 在光激发时由非荧光分子耗散的热。以这种方式，非荧光和潜在的 即使是弱吸收的光谱特征，例如那些诊断金属配位环境的特征， 可以解析出loenzyme。在第二种互补几何形状中，我们利用了珀塞尔效应， 可以显著提高小模体积和高质量光学微腔中的散射率 因素虽然Sers基本上需要与等离子体表面的货车范德华接触，但微腔内 增强剂可以在距离电介质表面高达~100 nm的距离处起作用，使其适合于探测生物 分子没有明显的扰动。我们现在已经展示了这两个战略背后的核心概念- 埃及人在具体目标1-3中，我们将在线评估三种新的微腔系统，这些系统承诺 显著提高了我们的测量能力，足以为生物医学应用奠定坚实的基础：平面 氮化硅环形谐振器（SA 1）、光纤法布里-珀罗微腔（SA 2）和氮化硅纳米梁 （SA3）。在所有情况下，我们将对一系列越来越具有挑战性的粒子和分子进行光谱学研究， 推动对单一工作金属酶的监测。支持计算表明，这些 新的共振器几何结构将使我们的分子信号增加几个数量级。我们的长期目标 是将一种新的、高信息量的、甚至是破坏性的生物物理技术应用于生物分子 了解它们是如何运作的，如何随时间变化，如何受到监管，如何失败。