Thermodynamic and Directed Physical Characterization of Bacteriophage Lambda Capsid Maturation

Abstract

All viruses undergo a multistep developmental process to assemble a mature virus. An essential step in the assembly of complex double-stranded DNA viruses is packaging the viral genome into a pre-formed procapsid shell. In bacteriophage , packaging of ~15 kb of DNA triggers a dramatic conformational change that expands the shell and increases the capsid volume two fold; this is a common feature in most dsDNA viruses. It has been recently demonstrated that expansion of the lambda procapsid is reversible and I have characterized the thermodynamic features of the transition. The data indicate that significant hydrophobic surface area is exposed in the expanded shell. It has been further shown that the gpD decoration protein adds to the expanded capsid lattice to stabilize the shell. GpD is a monomer in solution but self-assembles as a trimer spike at the three-fold verticies of the icosahedral capsid. Addition of gpD to the expanded capsid surface stabilizes the capsid from both external as well as internal forces. I propose that the hydrophobic patches exposed in the expanded capsid shell serve to nucleate gpD oligomerization at the capsid surface. I also propose that there are three additional non-covalent interactions that play important roles in stabilizing the expanded capsid from extreme internal pressure as DNA packaging is completed. Here I examine those interactions in detail along with gpD trimerization at the capsid surface using defined in vitro biochemical assay systems. The results of this thesis provide insight into the complex nature and importance of capsid maturation for bacteriophage lambda that are generalizable to all of the complex dsDNA viruses, both prokaryotic and eukaryotic.