title: Alterations in catalytic activity and virus maturation produced by mutation of the conserved histidine residues of herpes simplex virus type 1 protease.
abstract: Mutant herpes simplex virus type 1 (HSV-1) viruses were constructed to characterize the roles of the conserved histidine residues (H61 and H148) of HSV-1 protease in the regulation of catalytic activity and virus maturation. Viruses containing mutations at H61 (H61V-V711, H61Y-V715, and H61A-V730) were unable to grow on Vero cells. These mutant viruses could process neither Pra to N0 nor ICP-35cd to ICP-35ef. Transmission electron microscopy studies of H61A-V730-infected Vero cells indicated that capsid maturation is arrested at a state characterized by the predominance of large symmetrical arrays of B capsids within the nucleus. Two mutations at H148 (in viruses H148A-V712 and H148E-V728) gave rise to mutant viruses that grew with a small-plaque phenotype; one of the viruses, H148E-V728, was particularly attenuated when grown at a low multiplicity of infection. The rate of processing of Pra to N0 in infected Vero cells increased in the order H148A-V712 < H148E-V728 < parental strain HSV-1-V731. The observation that H148A-V712 processes Pra to N0 and ICP-35cd to ICP-35ef, whereas H61A does not, establishes H61 as the catalytically essential conserved His assuming that HSV-1 protease, like other serine proteases, utilizes an active-site histidine residue in catalysis. Two of the mutations at H148 (viruses H148K-V729 and H148Y-V716) produced nonviable viruses. H148K-V729 processed neither Pra to N0 nor ICP-35cd to ICP-35ef, whereas H148Y-V716 processed Pra to N0 but did not process ICP-35cd to ICP-35ef. The range of phenotypes observed with the H148 mutant viruses suggests that residue 148 of the HSV-1 protease is a determinant of virus growth rate and viability because of its effects on the activity of the protease and/or the role of the protease domain in capsid assembly and DNA packaging.
authors.affiliation: Department of Biological Chemistry, Merck Research Laboratories, West Point, Pennsylvania 19486, USA.
authors.forename: R B
authors.initials: RB
authors.lastname: Register
authors.forename: J A
authors.initials: JA
authors.lastname: Shafer
mesh_terms: D019943:Amino Acid Substitution; D000818:Animals; D002213:Capsid / Q000378:metabolism*; D002384:Catalysis; D002522:Chlorocebus aethiops; D018259:Herpesvirus 1, Human / Q000201:enzymology*; D006639:Histidine; D008854:Microscopy, Electron; D017354:Point Mutation; D012697:Serine Endopeptidases / Q000378:metabolism*; D013379:Substrate Specificity; D014709:Vero Cells; D014764:Viral Proteins; D014779:Virus Replication
chemical_list: D014764:Viral Proteins; C079287:scaffold protein, Herpes simplex virus-1; D006639:Histidine; D012697:Serine Endopeptidases