Application of antibody to synthetic peptides for characterization of the intact and truncated alpha 22 protein specified by herpes simplex virus 1 and the R325 alpha 22- deletion mutant.
The alpha 22 protein is one of five proteins synthesized immediately after infection of permissive cells with herpes simplex virus 1 and 2 (HSV-1 and HSV-2). On the basis of the reported nucleotide sequence of the HSV-1 gene, we synthesized two peptides containing the predicted amino acids 12 through 23 (12 residues) and 21 through 36 (16 residues) in two hydrophilic domains near the N terminus of the protein. Rabbit antisera made against these peptides were then used to characterize the alpha 22 protein made by wild-type HSV-1(F) strain and by an HSV-1 mutant, R325, carrying a 500-base-pair deletion within the coding domain of the gene. The results were as follows. (i) Both antisera reacted with HSV-1(F) alpha 22 protein in lysates electrophoretically separated in denaturing polyacrylamide gels and electrically transferred to a nitrocellulose sheet; neither antiserum reacted with the corresponding HSV-2 protein. The protein accumulated at 34 and 39 degrees C in the nucleus of infected permissive HEp-2 and baby hamster kidney (BHK) cells. The protein formed at least five spots differing in charge, mobility, and extent of phosphorylation on two-dimensional electrophoretic separation. (ii) The antisera reacted with a truncated nuclear protein (33,700 apparent molecular weight) in permissive HEp-2 and restrictive BHK cells infected with R325 and incubated at 39 degrees C but not at 34 degrees C. The truncated protein represents, therefore, the product of the undeleted 5' domain of the alpha 22 gene in R325. (iii) The presence of identical as well as slower migrating, reactive proteins in infected BHK cell lysates indicated that wild-type and truncated alpha 22 proteins are processed differently in BHK and HEp-2 cells.
Ackermann
M
M
Sarmiento
M
M
Roizman
B
B
D000595:Amino Acid Sequence; D000818:Animals; D000918:Antibody Specificity; D002451:Cell Compartmentation; D002460:Cell Line; D002872:Chromosome Deletion; D006224:Cricetinae; D004274:DNA, Recombinant; D006801:Humans; D009154:Mutation; D010446:Peptide Fragments / Q000138:chemical synthesis / Q000276:immunology; D010766:Phosphorylation; D011817:Rabbits; D018139:Simplexvirus / Q000235:genetics* / Q000276:immunology; D012995:Solubility; D014764:Viral Proteins / Q000235:genetics* / Q000276:immunology
D004274:DNA, Recombinant; D010446:Peptide Fragments; D014764:Viral Proteins