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

向卓越的过渡项目将支持首席研究人员的专业发展，以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)。在该项目的学习阶段完成后，这些新技术将被应用于深入了解负责获取结构和维持大型复杂蛋白质结构的基本生物过程。目前对大型蛋白质的这些过程知之甚少，但它们对蛋白质正确的生物学功能至关重要；功能障碍可能会导致一些严重的疾病，包括帕金森氏症等神经退行性疾病。SP Cryo-EM最近给结构生物学带来了革命性的变化，因此该项目有可能在迄今无法达到的三维细节水平上促进蛋白质结构获取和恢复过程的基础知识，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。该项目包括对SP Cryo-EM进行密集的理论和实验培训，并将从专业上提升一名高级研究员和若干博士后、博士后和本科生。该项目将促进使用低温电子显微镜的新的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。这一项目将使首席调查员能够将研究能力大大扩展到高分辨率(近原子)低温电子显微镜、蛋白质折叠的结构研究和通过伴侣进行蛋白质结构恢复机制等领域。这一转变将使人们能够对大分子内部和之间复杂的原子间相互作用有新的见解，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。将原子力显微镜等分子级分辨率技术与低温电子显微镜的高分辨率结构方法相结合，将能够详细描述蛋白质生物合成与超大型蛋白质(如锚蛋白和萤火虫荧光素酶)折叠路径之间的关系，这些关系以前无法在近本地条件下以原子分辨率进行研究。该奖项反映了NSF的法定使命，并通过使用基金会的智力优势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.