A Domain of Herpes Simplex Virus pU L 33 Required To Release Monomeric Viral Genomes from Cleaved Concatemeric DNA.

Monomeric herpesvirus DNA is cleaved from concatemers and inserted into preformed capsids through the actions of the viral terminase. The terminase of herpes simplex virus (HSV) is composed of three subunits encoded by UL 15, UL 28, and UL 33. The UL 33-encoded protein (pUL 33) interacts with pUL 28, but its precise role in the DNA cleavage and packaging reaction is unclear. To investigate the function of pUL 33, we generated a panel of recombinant viruses with either deletions or substitutions in the most conserved regions of UL 33 using a bacterial artificial chromosome system. Deletion of 11 amino acids (residues 50 to 60 or residues 110 to 120) precluded viral replication, whereas the truncation of the last 10 amino acids from the pUL 33 C terminus did not affect viral replication or the interaction of pUL 33 with pUL 28. Mutations that replaced the lysine at codon 110 and the arginine at codon 111 with alanine codons failed to replicate, and the pUL 33 mutant interacted with pUL 28 less efficiently. Interestingly, genomic termini of the large (L) and small (S) components were detected readily in cells infected with these mutants, indicating that concatemeric DNA was cleaved efficiently. However, the release of monomeric genomes as assessed by pulsed-field gel electrophoresis was greatly diminished, and DNA-containing capsids were not observed. These results suggest that pUL 33 is necessary for one of the two viral DNA cleavage events required to release individual genomes from concatemeric viral DNA. IMPORTANCE This paper shows a role for pUL 33 in one of the two DNA cleavage events required to release monomeric genomes from concatemeric viral DNA. This is the first time that such a phenotype has been observed and is the first identification of a function of this protein relevant to DNA packaging other than its interaction with other terminase components.