Expression of cell-associated and secreted forms of herpes simplex virus type 1 glycoprotein gB in mammalian cells.
The gene for glycoprotein gB1 of herpes simplex virus type 1 strain Patton was expressed in stable Chinese hamster ovary cell lines. Expression vectors containing the dihydrofolate reductase (dhfr) cDNA plus the complete gB1 gene or a truncated gene lacking the 194 carboxyl-terminal amino acids of gB1 were transfected into CHO DHFR-deficient cells. Radioimmunoprecipitation demonstrated that the complete gB1 protein expressed in CHO cell lines was cell associated, whereas the truncated protein was secreted from the cells due to deletion of the transmembrane and C-terminal domains of gB1. Cells expressing the truncated gB1 protein were subjected to stepwise methotrexate selection, and a cell line was isolated in which the gB1 gene copy number had been amplified 10-fold and the level of expression of gB1 had increased over 60-fold. The truncated gB1 protein was purified from medium conditioned by the amplified cell line. N-terminal amino acid sequence analysis of this purified protein identified the signal peptide cleavage site and predicted the cleavage of a 30-amino-acid signal sequence from the primary protein. The immunogenicity of the truncated gB1 protein was also tested in mice, and high levels of antibody and protection from virus challenge were observed.
D000595:Amino Acid Sequence; D000818:Animals; D002460:Cell Line; D006224:Cricetinae; D003412:Cricetulus; D004247:DNA / Q000378:metabolism; D004262:DNA Restriction Enzymes; D005260:Female; D005796:Genes; D005814:Genes, Viral; D010053:Ovary; D018139:Simplexvirus / Q000235:genetics*; D013762:Tetrahydrofolate Dehydrogenase / Q000235:genetics; D014162:Transfection; D014759:Viral Envelope Proteins / Q000235:genetics* / Q000378:metabolism
D014759:Viral Envelope Proteins; C040495:glycoprotein B, Simplexvirus; D004247:DNA; D013762:Tetrahydrofolate Dehydrogenase; D004262:DNA Restriction Enzymes