In bacteriophage T7 as well as many other dsDNA phages and some animal viruses, scaffolding protein is essential for the accurate formation of the metastable precursor particle (prohead) and thus it is a vital aspect of the viral infection cycle. When purified by anion exchange and gel filtration chromatography, the T7 scaffolding protein (gp9) exists as an extended monomer in solution. These monomers of gp9 associate with other scaffolding monomers to form the extended filaments in T7 procapsids visualized by electron tomography. These filaments interact with the negatively charged inner surface of the procapsid at unique sites, probably via the extended positively charged C-terminus of gp9. Scaffolding protein, via these interactions, facilitates isometric capsid assembly by helping to define the proper curvature of the viral capsid. Consequently, scaffolding-mediated viral assembly does not require a rigid structural network, nor does it require a stoichiometric amount of gp9 to be present in each procapsid. The observed flexibility of the scaffolding network, association of scaffolding filaments with the core connector complex, as well as the variable copy number of gp9 per particle suggests an assembly mechanism where capsid formation is nucleated around the core/connector complex. In such a mechanism there is more than one path to a single end - production of the correctly formed prohead particles that are required for T7 phage maturation.
|Advisor:||Beckett, Dorothy, Steven, Alasdair C.|
|Commitee:||Hendrix, Roger W., Lorimer, George H., Mount, Stephen M.|
|School:||University of Maryland, College Park|
|School Location:||United States -- Maryland|
|Source:||DAI-B 70/06, Dissertation Abstracts International|
|Keywords:||Bacteriophage T7, Electron tomography, Procapsid, Scaffolding proteins, Virus assembly, dsDNA phage|
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