An unexpected twist in viral capsid maturation

Abstract

Lambda-like dsDNA bacteriophage undergo massive conformational changes in their capsid shell during the packaging of their viral genomes. Capsid shells are complex organizations of hundreds of protein subunits that assemble into intricate quaternary complexes that ultimately are able to withstand over 50 atm. of pressure during genome packaging 1 . The extensive integration between subunits in capsids is unlikely to form in a single assembly step, therefore requiring formation of an intermediate complex, termed a procapsid, from which individual subunits can undergo the necessary refolding and structural rearrangements needed to transition to the more stable capsid. Though various mature capsids have been characterized at atomic resolution, no such procapsid structure is available for a dsDNA virus or bacteriophage that undergoes large scale conformational changes. We present a procapsid x-ray structure at 3.65Å resolution, termed Prohead II, of the lambda like bacteriophage HK97, whose mature capsid structure was previously solved to 3.44 Å 2 . A comparison of the two largely different capsid forms has unveiled an unprecedented expansion mechanism that describes the transition. Crystallographic and Hydrogen/Deuterium exchange data presented here demonstrates that the subunit tertiary structures are significantly different between the two states, with twisting and bending motions occurring in both helical and β-sheet regions. We have also discovered conserved subunit interactions at each 3-fold of the virus capsid, from which capsid subunits maintain their integrity during refolding, facilitating the rotational and translational motions of maturation. Calormetric data of a closely related bacteriophage, P22, showed that capsid maturation was an exothermic process that resulted in a release of 90KJ/mol of energy 3 . We propose the major tertiary changes presented in this study reveal a structural basis for an exothermic maturation process likely present in many dsDNA Bacteriophage and possibly viruses such as Herpes which share the HK97 subunit fold 4 . Main TextHK97 is a favorable system for studying capsid maturation because capsid particles can be assembled in E. coli from the expression of just two viral gene products, gp4 (protease) and gp5 (capsid subunit), and maturation can be triggered and analyzed in vitro 2,5-8 . The 384-residue gp5 subunits assemble into hexameric and pentameric oligomers, termed capsomers, that first assemble to form the Prohead I capsid (P-I). The 17.7-Megadalton, T=7 laevo icosahedral particle contains 12 pentamers and 60 hexamers and encapsidates approximately 60 copies of the gp4 protease 9-11 . Prohead I particles can be made with either a defective protease or without protease and can be disassembled in vitro into free capsomers and than reassembled when exposed to specific chemical treatments 12 . When active gp4 is present, the particles spontaneously mature to the Prohead II (P-II) form following digestion of residues 2-103 from all subunits. The proteolytic fragment...

Molecular Rearrangements Involved in the Capsid Shell Maturation of Bacteriophage T7

“…3A), which makes them more parallel to the shell surface. A similar relative movement between the A-and P-domains has previously been found in the maturation of HK97 (46) and P22 (20). Monomers were rearranged in the capsomers of the head relative to the prohead.…”

Visualization of uncorrelated, tandem symmetry mismatches in the internal genome packaging apparatus of bacteriophage T7

“…S2B), as shown before in the procapsid structures of T7 (19,26) and other tailed dsDNA phages (10,27). In the icosahedral reconstruction, all of the nonicosahedral features, including the scaffolding protein (gp9) inside the gp10 shell and the portal/core stack complex, are smeared.…”

Orientational phase transitions and the assembly of viral capsids

“…However, HK97 is exceptional as it does not form on a precursor scaffold. Instead, the capsid proteins are linked by an interconnected net of ∆ groups below the actual capsid [39]. This net is believed to play the role of a scaffold.…”