| Description |
During retroviral maturation, the viral CA protein oligomerizes to form a closed capsid that surrounds the viral genome. Interestingly, mature capsid morphologies vary dramatically across the different genera of retroviridae. Specifically, capsids can be conical, spherical, or cylindrical. Despite these morphological differences, however, the tertiary structure of CA proteins is conserved across all genera, suggesting that there may be commonalities in the structure of all capsids. This dissertation reports biochemical and structural studies of retroviral CA assemblies formed in vitro, with the goal of understanding the structure and conservation of retroviral capsids. Helical assemblies made from HIV-1 CA are composed of hexamers. This work led us to propose that the conical HIV-1 capsid could be built on a hexameric lattice closed by asymmetrically introducing twelve pentameric defects. A corollary of our model is that within these conical assemblies, only five cone angles are allowed. Therefore, we developed a method for producing HIV-1 CA conical assemblies in vitro, and measurements of ?1,000 of these synthetic cones revealed that the predicted angles are preferentially populated. High-resolution studies of the HIV-1 CA protein have been combined with helical reconstructions of HIV-1 CA to create a pseudoatomic model of the viral capsid. CA interfaces present in the model are predicted to be important for HIV-1 assembly. Indeed, alanine scanning mutagenesis demonstrated that these interfaces are important for CA cylinder assembly in vitro. Interestingly, this study also revealed a CA mutant (R18A), which could assemble into spheres, cylinders and cones (the three capsid morphologies seen within authentic virions). Finally, we have proposed that all retroviral capsids are composed of hexameric arrays of CA protein similar to those defined for HIV-1, and that the distinct morphologies of retroviral capsids reflect different distributions of the pentameric declinations that allow the structures to close. Therefore, we determined the three dimensional structure of MMuLV CA crystallized on a lipid monolayer and found that M-MuLV also assembles into hexamers similar to those reported for HIV-1. In summary, this dissertation proposes a general model for retroviral assembly, wherein all retroviral capsids are built on similar hexameric lattices closed by pentameric defects. |
