Macromolecular assemblies are involved in nearly every process in the cell, from motility to signaling. Typically, such assemblies are composed of a variety of protein and organized into intricate machines that undergo dynamic and transient shifts in their organization and composition. As such, understanding macromolecular assemblies is paramount for increasing our knowledge about basic cellular events and discovering targets for disease prevention. While electron cryo-microscopy (cryo-EM) is capable of imaging these large and often difficult to study assemblies, limitations currently hinder the ability to atomic resolutions, at which point these structures can reveal the intricate sets of interactions and cordination of complex chemistry. State-of-the-art cryo-EM imaging has recently achieved near-atomic resolutions (3.5-5?), such that basic models for all or parts of an entire macromolecular assembly could be constructed. These """"""""first- approach"""""""" models describe the organization and basic fold of the protein constituents, but fall short of describing the necessary atomic-level detail features of macromolecular assemblies that is critical for designing effective measures for disease prevention and improving human health. Motivated by the need for new tools to push towards atomic resolution structures in cryo-EM, we propose developing a comprehensive macromolecular refinement and modeling protocol. Our ultimate goal is to provide the community with a new technique to push near-atomic resolution structures to atomic resolutions such that full atomic models can be built directly from a density map. To accomplish this goal, we will develop a novel automated method for generating simple structural models that, in turn can be used to refine and improve resolution in a cryo-EM density map. From this, it will not only be possible to construct fill atomic models, but also to model the structural dynamics captured by cryo-EM imaging. We believe this work will revolutionize the field of cryo-EM, as well as lead to significant leaps in our understanding of all biological processes.

Public Health Relevance

Macromolecular assemblies play critical roles in nearly every biological process and, as such, understanding their structure and function is critical to human health. While limited in resolution, electron cryo-microscopy (cryo-EM) is one of the few techniques capable of visualizing such assemblies in near-native conditions. In this proposal, we aim to enhance the resolution and build complete atomic models for macromolecular assemblies imaged with cryo-EM using a novel model building and refinement strategy, leading to a better understanding of cellular processes and disease prevention.

National Institute of Health (NIH)
National Institute of General Medical Sciences (NIGMS)
Exploratory/Developmental Grants (R21)
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Macromolecular Structure and Function D Study Section (MSFD)
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Flicker, Paula F
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Baylor College of Medicine
Schools of Medicine
United States
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Dou, Hang; Burrows, Derek W; Baker, Matthew L et al. (2017) Flexible Fitting of Atomic Models into Cryo-EM Density Maps Guided by Helix Correspondences. Biophys J 112:2479-2493
Chen, Muyuan; Baldwin, Philip R; Ludtke, Steven J et al. (2016) De Novo modeling in cryo-EM density maps with Pathwalking. J Struct Biol 196:289-298
Fan, Guizhen; Baker, Matthew L; Wang, Zhao et al. (2015) Gating machinery of InsP3R channels revealed by electron cryomicroscopy. Nature 527:336-41