Optical Control of Protein Activity in Live Cells by Plasmon Assisted Light Inactivation

通过等离激元辅助光灭活对活细胞中蛋白质活性的光学控制

• 批准号: 10698186
• 负责人: Zhenpeng Qin
• 金额: $ 38.25万
• 依托单位: UNIVERSITY OF TEXAS DALLAS
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-08-01 至 2024-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10698186
• 关键词: Address; Biological Assay; Biology; Cells; Cellular biology; Collaborations; Communities; Development; Diagnostic Procedure; G Protein-Coupled Receptor Signaling; G-Protein-Coupled Receptors; GTPase-Activating Proteins; Goals; Gold; Heating; Investigation; Laboratory Research; Lasers; Light; Lipids; Location; Measures; Mediating; Membrane; Methods; Modeling; Modern Medicine; Molecular; Neuropeptide Receptor; Neuropeptides; Optical Methods; Optics; Outcome; PAR-2 Receptor; Physiologic pulse; Process; Protein Conformation; Proteins; Recording of previous events; Research; Signal Transduction; Spectrum Analysis; Surface; Techniques; Temperature; Testing; Time; Work; biological systems; chronic pain; extracellular; innovation; insight; interest; nano; nanomaterials; nanometer; nanoparticle; nanosecond; nanovesicle; novel; plasmonics; programs; receptor; response; success; temperature jump; tool; trafficking

Abstract Optical tools have unparalleled spatial and temporal precision and have been instrumental to better understand various processes in modern medicine and biology. The overall goal of my research laboratory is to understand the laser-plasmonic nanoparticle interactions and its effects at the interface between biological systems and nanomaterials. Specifically, experimental techniques and methods have been developed to understand the effects of nanoparticle plasmonic heating on proteins and lipids immediately next to the nanoparticle. This has led to new enabling tools for optical protein manipulation and photosensitive nanovesicles for molecular uncaging, as well as innovative diagnostic methods. This proposed research focus on the development of optical control of protein activity in live cells, namely plasmon-assisted light inactivation (PALI). PALI is based on pulsed laser heating (nanosecond) of plasmonic nanoparticles, and its thermally confined heating to unfold and denature surrounding proteins within a few nanometers of nanoparticle surface. Thus, PALI also effectively acts a unique nano temperature-jump (T-jump), an innovative experimental platform to address a gap for protein unfolding investigations. In the next five years, I plan to develop my research program in these two directions. Firstly, I will focus on developing this new optical tool to manipulate protein activity in live cells with emphasis on G-protein coupled receptors (GPCR), an important and diverse class of membrane receptors that mediate extracellular to intracellular signaling. This encompasses a systematic approach to understand the interaction and trafficking of nanoparticles with GPCR, the cellular responses of PALI on GPCR signaling, and finally the applicability of PALI on other GPCRs. I will primarily use a specific GPCR, protease activated receptor 2 (PAR2) that is important for chronic pain, as a working model. To test for other GPCRs, I will test GPCRs for neuropeptides, which are synergistic with our efforts to create neuropeptide photosensitive nanovesicles. Secondly, I will concentrate on the characterization of the nano T-jump by addressing two fundamental questions: (1) can the nanoparticle temperature be directly measured during pulsed laser heating or after a short delay? (2) How does the protein unfold under nano T-jump? These involves our existing collaborations with the Argonne National Lab to probe the gold lattice expansion using advanced spectroscopy, and various structural and functional assays to measure the protein unfolding and inactivation due to the nano T-jump. By the end of the five years, I anticipate solving important technical challenges to demonstrate the use of PALI to manipulate protein activity in live cells through GPCRs, and obtain a clear understanding of the temperature history and protein responses with the innovative nano T-jump platform. These outcomes would generate interest to the broad research community and enable others to tackle important challenges in cell biology using PALI.

抽象的 光学工具具有无与伦比的空间和时间精度，并且有助于更好地理解 现代医学和生物学的各种过程。我的研究实验室的总体目标是 了解生物学之间的激光质量纳米粒子相互作用及其在界面上的影响 系统和纳米材料。具体而言，已经开发了实验技术和方法 了解纳米颗粒等离激元加热对紧邻蛋白质和脂质的影响 纳米颗粒。这导致了用于光学蛋白质操纵和光敏的新的促成工具 用于分子渗透的纳米孢子以及创新的诊断方法。这个拟议的研究重点 关于活细胞中蛋白质活性的光学控制，即等离子辅助光失活 （帕利）。 Pali基于等离子纳米颗粒的脉冲激光加热（纳米秒）及其热量 将加热限制为展开，并在纳米颗粒表面的几纳米内蛋白质变性。 因此，帕利还有效地作用独特的纳米温度跳跃（T-JUMP），这是一个创新的实验平台 解决蛋白质展开研究的差距。在接下来的五年中，我计划发展我的研究 在这两个方向上进行程序。首先，我将专注于开发这种新的光学工具来操纵蛋白质 活细胞的活性，重点是G蛋白偶联受体（GPCR），这是一类重要而多样的类别 介导细胞外信号传导的膜受体。这包括系统 了解纳米颗粒与GPCR的相互作用和运输的方法， Pali在GPCR信号传导上，最后是Pali在其他GPCR上的适用性。我将主要使用特定的 GPCR，蛋白酶激活受体2（PAR2），对于慢性疼痛作为工作模型很重要。测试 其他GPCR，我将测试GPCR的神经肽，这与我们创建神经肽的努力协同作用 光敏性纳米层。其次，我将专注于纳米T-突变的表征 解决两个基本问题：（1）可以直接测量纳米颗粒温度 脉冲激光加热还是短延迟后？ （2）蛋白在纳米T-Jump下如何展开？这些 涉及我们与Argonne National Lab的现有合作，以使用 晚期光谱法以及各种结构和功能测定法，以测量蛋白质展开和 由于纳米T型引起的失活。到五年结束时，我预计要解决重要的技术 挑战以证明使用PALI通过GPCR操纵活细胞中的蛋白质活性，并 通过创新的纳米T-大小对温度病史和蛋白质反应有清晰的了解 平台。这些结果将引起广泛的研究社区的兴趣，并使其他人能够 解决使用Pali的细胞生物学的重要挑战。