The most general goal of this project, ever since its inception in 1972, has been to gather the structural data for realistic "molecular movies" of virus assembly, entry, and uncoating. In so doing, we seek to establish mechanisms for quite general processes, such as membrane fusion and perforation, as well as to lay the groundwork for discovering novel classes of antiviral therapeutics and for designing new vaccines. During the three-and-one-half years covered by this Progress Report, we have combined x-ray crystallography with electron cryo-microscopy (cryoEM) to analyze the organization of reovirus and rotavirus particles and to study the conformational changes in their penetration proteins (u1 and VP5*, respectively) that lead to membrane perforation and translocation into the cytoplasm. We have determined the structure of the rotavirus dsRNA-dependent RNA polymerase, VP1, and compared it with the related polymerase from reovirus. We are engaged in an on-going effort to advance the resolution of cryoEM-generated 3-D image reconstructions of human papillomavirus-like particles (HPV VLPs). We have also analyzed the conformational states of the SARS coronavirus spike protein and determined the structure of its receptor- binding domain bound with a soluble form of its receptor (ACE2). In the coming period, we aim to complete the HPV VLP effort;to reconstitute a full atomic model of the rotavirus particle, by combining cryoEM and x- ray structures;to study the molecular mechanism of rotavirus budding and maturation in the ER;to build upon current results to work out a full picture of reovirus penetration;to do likewise for rotavirus;and to continue work on viral polymerases, by initiating structural studies of vesicular stomatitis virus RNA- dependent RNA polymerase.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Method to Extend Research in Time (MERIT) Award (R37)
Project #
5R37CA013202-41
Application #
8251202
Study Section
Special Emphasis Panel (NSS)
Program Officer
Knowlton, John R
Project Start
1975-06-01
Project End
2013-04-30
Budget Start
2012-05-01
Budget End
2013-04-30
Support Year
41
Fiscal Year
2012
Total Cost
$378,081
Indirect Cost
$160,793
Name
Children's Hospital Boston
Department
Type
DUNS #
076593722
City
Boston
State
MA
Country
United States
Zip Code
02115
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Abraham, Jonathan; Corbett, Kevin D; Farzan, Michael et al. (2010) Structural basis for receptor recognition by New World hemorrhagic fever arenaviruses. Nat Struct Mol Biol 17:438-44
Chen, James Z; Settembre, Ethan C; Aoki, Scott T et al. (2009) Molecular interactions in rotavirus assembly and uncoating seen by high-resolution cryo-EM. Proc Natl Acad Sci U S A 106:10644-8
Aoki, Scott T; Settembre, Ethan C; Trask, Shane D et al. (2009) Structure of rotavirus outer-layer protein VP7 bound with a neutralizing Fab. Science 324:1444-7

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