Helical polymers are formed by many proteins found in bacterial, archaeal and eukaryotic cells, and can also be present as viral capsids, nucleocapsids and tails. In certain cases most of the protein found in a cell is in the form of a helical polymer, so methods to study the structure and dynamics of such polymers have great general interest. Particular helical complexes, such as the pili and flagellar filaments of pathogenic bacteria, have a very immediate relevance to human disease. We propose to further develop, extend, apply and support new methods for the three-dimensional reconstruction of such polymers from electron microscopic images. Our work in this area has already had a large impact on a number of NIH-supported projects from many laboratories, but we are now at a transition point where the potential for achieving high- resolution structures from numerous samples is now quite high. Given the imminent arrival of a Titan Krios TEM we need support to further develop our Iterative Helical Real Space Reconstruction approach and optimize the processing of large numbers of high-resolution images of polymers. Effort will be invested in using new algorithms for alignment and reconstruction, as well as in developing methods that make use of prior knowledge about the spatial relations between different segments that have been cut from the same filament. All of the development work will use samples that have great interest to a large community and that have a direct relation to human health. These range from bacterial pili to viral capsids to the protein product of an oncogene. We have established that processing of such images can scale with the number of processors, so effort will be invested in making these programs easy to use on relatively inexpensive and commercially-available clusters. Tools for detecting potential ambiguities in helical symmetry will be developed, but the main tool will be bringing the resolution of helical reconstructions to the point where secondary structure can be recognized. At this resolution these ambiguities disappear.

Public Health Relevance

Many bacteria that cause human diseases require helical polymers on their surface for both attachment and motility. Helical viruses also exist, as do helical polymers formed by proteins from HIV and other pathogenic viruses. Developing improved methods to study the structure of helical polymers will thus have a direct impact on understanding and preventing many diseases.

Agency
National Institute of Health (NIH)
Institute
National Institute of Biomedical Imaging and Bioengineering (NIBIB)
Type
Research Project (R01)
Project #
5R01EB001567-11
Application #
8427216
Study Section
Enabling Bioanalytical and Imaging Technologies Study Section (EBIT)
Program Officer
Pai, Vinay Manjunath
Project Start
2003-02-01
Project End
2015-01-31
Budget Start
2013-02-01
Budget End
2014-01-31
Support Year
11
Fiscal Year
2013
Total Cost
$320,282
Indirect Cost
$108,107
Name
University of Virginia
Department
Biochemistry
Type
Schools of Medicine
DUNS #
065391526
City
Charlottesville
State
VA
Country
United States
Zip Code
22904
Egelman, Edward H (2016) The Current Revolution in Cryo-EM. Biophys J 110:1008-12
Kolappan, Subramania; Coureuil, Mathieu; Yu, Xiong et al. (2016) Structure of the Neisseria meningitidis Type IV pilus. Nat Commun 7:13015
Hospenthal, Manuela K; Redzej, Adam; Dodson, Karen et al. (2016) Structure of a Chaperone-Usher Pilus Reveals the Molecular Basis of Rod Uncoiling. Cell 164:269-78
Braun, Tatjana; Vos, Matthijn R; Kalisman, Nir et al. (2016) Archaeal flagellin combines a bacterial type IV pilin domain with an Ig-like domain. Proc Natl Acad Sci U S A 113:10352-7
Fu, Tian-Min; Li, Yang; Lu, Alvin et al. (2016) Cryo-EM Structure of Caspase-8 Tandem DED Filament Reveals Assembly and Regulation Mechanisms of the Death-Inducing Signaling Complex. Mol Cell 64:236-250
Wang, Ray Yu-Ruei; Kudryashev, Mikhail; Li, Xueming et al. (2015) De novo protein structure determination from near-atomic-resolution cryo-EM maps. Nat Methods 12:335-8
DiMaio, Frank; Song, Yifan; Li, Xueming et al. (2015) Atomic-accuracy models from 4.5-Ã… cryo-electron microscopy data with density-guided iterative local refinement. Nat Methods 12:361-5
Egelman, E H; Xu, C; DiMaio, F et al. (2015) Structural plasticity of helical nanotubes based on coiled-coil assemblies. Structure 23:280-9
Kudryashev, Mikhail; Wang, Ray Yu-Ruei; Brackmann, Maximilian et al. (2015) Structure of the type VI secretion system contractile sheath. Cell 160:952-62
Egelman, Edward H (2015) Three-dimensional reconstruction of helical polymers. Arch Biochem Biophys 581:54-8

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