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

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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.