Architect of Virus Assembly: the Portal Protein Nucleates Procapsid Assembly in Bacteriophage P22 (original) (raw)

2019, Journal of Virology

Tailed double-stranded DNA (dsDNA) bacteriophages, herpesviruses, and adenoviruses package their genetic material into a precursor capsid through a dodecameric ring complex called the portal protein, which is located at a unique 5-fold vertex. In several phages and viruses, including T4, ⌽29, and herpes simplex virus 1 (HSV-1), the portal forms a nucleation complex with scaffolding proteins (SPs) to initiate procapsid (PC) assembly, thereby ensuring incorporation of only one portal ring per capsid. However, for bacteriophage P22, the role of its portal protein in initiation of procapsid assembly is unclear. We have developed an in vitro P22 assembly assay where portal protein is coassembled into procapsid-like particles (PLPs). Scaffolding protein also catalyzes oligomerization of monomeric portal protein into dodecameric rings, possibly forming a scaffolding protein-portal protein nucleation complex that results in one portal ring per P22 procapsid. Here, we present evidence substantiating that the P22 portal protein, similarly to those of other dsDNA viruses, can act as an assembly nucleator. The presence of the P22 portal protein is shown to increase the rate of particle assembly and contribute to proper morphology of the assembled particles. Our results highlight a key function of portal protein as an assembly initiator, a feature that is likely conserved among these classes of dsDNA viruses. IMPORTANCE The existence of a single portal ring is essential to the formation of infectious virions in the tailed double-stranded DNA (dsDNA) phages, herpesviruses, and adenoviruses and, as such, is a viable antiviral therapeutic target. How only one portal is selectively incorporated at a unique vertex is unclear. In many dsDNA viruses and phages, the portal protein acts as an assembly nucleator. However, early work on phage P22 assembly in vivo indicated that the portal protein did not function as a nucleator for procapsid (PC) assembly, leading to the suggestion that P22 uses a unique mechanism for portal incorporation. Here, we show that portal protein nucleates assembly of P22 procapsid-like particles (PLPs). Addition of portal rings to an assembly reaction increases the rate of formation and yield of particles and corrects improper particle morphology. Our data suggest that procapsid assembly may universally initiate with a nucleation complex composed minimally of portal and scaffolding proteins (SPs). KEYWORDS coat protein, connector, initiation complex, terminase D ouble-stranded DNA (dsDNA) tailed bacteriophages, herpesviruses, some dsDNA archaeal viruses, and adenoviruses have a pseudoicosahedral capsid with a portal protein incorporated at a unique 5-fold symmetry axis (1-3). The portal protein is a ring of 12 identical subunits constituting a channel for the active packaging and subsequent ejection of the dsDNA viral genome (4-8). The existence of a single portal ring is crucial to the formation of infectious virions. Evidence from dsDNA bacteriophages, such as T4, SPP1, and ⌽29, and from herpes simplex virus 1 (HSV-1), indicates that portal proteins play a key role in the nucleation and assembly of proper procapsids, thereby ensuring one portal ring per capsid (9-13).