Transition To Excellence: From Single-Molecule Force Spectroscopy to Single-Particle Cryogenic Electron Microscopy

这个向卓越过渡的项目将支持首席研究员的专业发展，学习尖端的高分辨率显微镜技术：单粒子低温电子显微镜（SP Cryo-EM）。在项目的学习阶段完成后，这些新技术将被应用于了解大型复杂蛋白质的结构获取和结构维持的基本生物过程。这些过程目前对大型蛋白质知之甚少，但它们对蛋白质的正确生物学功能至关重要；功能障碍可能导致许多严重的疾病，包括神经退行性疾病，如帕金森病。SP Cryo-EM最近彻底改变了结构生物学，因此该项目有可能在迄今为止无法达到的三维细节水平上为蛋白质结构获取和恢复过程的基本知识做出贡献，这将对理解蛋白质折叠，错误折叠和结构恢复机制非常有用。该项目涉及SP Cryo-EM的理论和实验培训，将专业培养一名高级研究员和一些博士后、博士和本科生。本项目将促进使用低温电子显微镜的新型高分辨率结构方法来表征蛋白质折叠途径，这是传统方法难以实现的。该项目将使首席研究员能够将研究能力扩展到高分辨率（近原子）低温电子显微结构研究领域，研究蛋白质折叠和伴侣蛋白结构恢复机制。这种转变将使人们对大分子内部和大分子之间复杂的原子间相互作用有了新的认识，这些相互作用决定了结构的形成和动力学，这对获得或恢复生物功能至关重要。分子水平分辨率技术（如原子力显微镜）与高分辨率低温电子显微镜结构方法的结合，将允许详细表征蛋白质生物合成和非常大的蛋白质（如锚蛋白和萤火虫荧光素酶）折叠途径之间的关系，这些蛋白质在接近天然条件下无法在原子分辨率下进行研究。该奖项反映了美国国家科学基金会的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。

This Transition to Excellence Project will support the Principal Investigator’s Professional Development to learn the cutting edge high-resolution microscopy technique: Single-Particle Cryogenic Electron Microscopy (SP Cryo-EM). After the learning phase of the Project is completed, these new techniques will be applied to gain insights into fundamental biological processes responsible for the acquisition of structure and for structural maintenance of large and complex proteins. These processes are currently poorly understood for large proteins, yet they are critical for proteins’ correct biological functions; malfunction may lead to a number of serious illnesses including neurodegenerative disorders such as Parkinson’s disease. SP Cryo-EM has recently revolutionized structural biology and therefore this project has a potential to contribute to the basic knowledge of protein structure acquisition and restoration processes at a hitherto unattainable level of three-dimensional detail, which will be extremely useful for understanding protein folding, misfolding and structural recovery mechanisms. The project involves intense theoretical and experimental training in SP Cryo-EM and will professionally advance one senior investigator and a number of postdoctoral, doctoral and undergraduate students. This project will promote the use of novel high resolution structural methods of cryogenic electron microscopy to characterize protein folding pathways, which have been difficult using traditional methods. This project will allow the Principal Investigator to undergo a major expansion of research capabilities into the field of high resolution (near atomic) cryogenic electron microcopy structural studies of protein folding and protein structure recovery mechanisms by chaperones. This transition will allow new insights into complex interatomic interactions inside and between macromolecules that determine structure formation and dynamics, which are critical for the acquisition or restoration of biological function. A combination of molecular-level resolution techniques such as atomic force microscopy with high resolution structural methods of cryogenic electron microscopy will allow detailed characterization of the relationships between protein biosynthesis and folding pathways for very large proteins such as ankyrins and firefly luciferase, which could not be studied before at atomic resolution under near native conditions.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.