Characterization of VP22 in herpes simplex virus-infected cells.
We examine biochemical characteristics of the herpes simplex virus (HSV) tegument protein VP22 by gel filtration, glycerol sedimentation, and chemical cross-linking experiments and use time course radiolabeling and immunoprecipitation assays to analyze its synthesis and interaction with other infected-cell proteins. VP22 was expressed as a delayed early protein with optimal synthesis requiring DNA replication. In immunoprecipitation assays, VP22 was found in association with several additional proteins including VP16 and a kinase activity likely to be that of UL13. Furthermore, in sizing chromatography experiments, VP22 was present in several higher-order complexes in infected cells. From gel filtration analysis the major form of VP22 migrated with a molecular mass of approximately 160 kDa, consistent with its presence as a tetramer, or a dimer complexed with other proteins, with a fraction of the protein migrating at larger molecular mass. In vitro-synthesized VP22 sedimented in a size range consistent with a mixture of tetramers and dimers. Short N- or C-terminal deletions resulted in migration almost exclusively as dimers, indicating that VP22, in the absence of additional virus-encoded proteins, could form higher-order assemblies, most likely tetramers, but that both N-and C-terminal determinants were required for stabilizing such assemblies. Consistent with this we found that isolated proteins encompassing either the N-terminal or C-terminal region of VP22 sedimented as dimers, and that the purified C-terminal domain could be cross-linked into dimeric structures. These results are discussed with regard to possible virus and host interactions involved in VP22 recruitment into virus particles.
Mouzakitis
G
G
MCRI, The Chart, Oxted, Surrey RH8 OTL, United Kingdom.
McLauchlan
John
J
Barreca
Cristina
C
Kueltzo
Lisa
L
O'Hare
P
P
D000595:Amino Acid Sequence; D002850:Chromatography, Gel; D004261:DNA Replication; D019281:Dimerization; D004591:Electrophoresis, Polyacrylamide Gel; D004847:Epithelial Cells / Q000821:virology*; D006367:HeLa Cells; D017405:Herpes Simplex Virus Protein Vmw65 / Q000378:metabolism; D018259:Herpesvirus 1, Human / Q000502:physiology*; D006801:Humans; D047468:Immunoprecipitation; D008969:Molecular Sequence Data; D008970:Molecular Weight; D011485:Protein Binding; D011494:Protein Kinases / Q000378:metabolism; D015678:Viral Structural Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000378:metabolism*
D017405:Herpes Simplex Virus Protein Vmw65; D015678:Viral Structural Proteins; C073413:herpes simplex virus type 1 protein VP22; D011494:Protein Kinases; C082380:UL13 protein, Simplexvirus