Skip to main content
PMC home page
Journal of Virology logoLink to Journal of Virology
. 1985 Oct;56(1):207–215. doi: 10.1128/jvi.56.1.207-215.1985

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.

M Ackermann, M Sarmiento, B Roizman
PMCID: PMC252507  PMID: 2993651

Abstract

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.

Full text

PDF
207

Images in this article

210Image
on p.210
210Image
on p.210
211Image
on p.211
212Image
on p.212
213Image
on p.213
213Image
on p.213

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Ackermann M., Braun D. K., Pereira L., Roizman B. Characterization of herpes simplex virus 1 alpha proteins 0, 4, and 27 with monoclonal antibodies. J Virol. 1984 Oct;52(1):108–118. doi: 10.1128/jvi.52.1.108-118.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Anderson K. P., Costa R. H., Holland L. E., Wagner E. K. Characterization of herpes simplex virus type 1 RNA present in the absence of de novo protein synthesis. J Virol. 1980 Apr;34(1):9–27. doi: 10.1128/jvi.34.1.9-27.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Batterson W., Roizman B. Characterization of the herpes simplex virion-associated factor responsible for the induction of alpha genes. J Virol. 1983 May;46(2):371–377. doi: 10.1128/jvi.46.2.371-377.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Braun D. K., Pereira L., Norrild B., Roizman B. Application of denatured, electrophoretically separated, and immobilized lysates of herpes simplex virus-infected cells for detection of monoclonal antibodies and for studies of the properties of viral proteins. J Virol. 1983 Apr;46(1):103–112. doi: 10.1128/jvi.46.1.103-112.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Campbell M. E., Palfreyman J. W., Preston C. M. Identification of herpes simplex virus DNA sequences which encode a trans-acting polypeptide responsible for stimulation of immediate early transcription. J Mol Biol. 1984 Nov 25;180(1):1–19. doi: 10.1016/0022-2836(84)90427-3. [DOI] [PubMed] [Google Scholar]
  6. Dixon R. A., Schaffer P. A. Fine-structure mapping and functional analysis of temperature-sensitive mutants in the gene encoding the herpes simplex virus type 1 immediate early protein VP175. J Virol. 1980 Oct;36(1):189–203. doi: 10.1128/jvi.36.1.189-203.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Ejercito P. M., Kieff E. D., Roizman B. Characterization of herpes simplex virus strains differing in their effects on social behaviour of infected cells. J Gen Virol. 1968 May;2(3):357–364. doi: 10.1099/0022-1317-2-3-357. [DOI] [PubMed] [Google Scholar]
  8. Honess R. W., Roizman B. Regulation of herpesvirus macromolecular synthesis. I. Cascade regulation of the synthesis of three groups of viral proteins. J Virol. 1974 Jul;14(1):8–19. doi: 10.1128/jvi.14.1.8-19.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Honess R. W., Roizman B. Regulation of herpesvirus macromolecular synthesis: sequential transition of polypeptide synthesis requires functional viral polypeptides. Proc Natl Acad Sci U S A. 1975 Apr;72(4):1276–1280. doi: 10.1073/pnas.72.4.1276. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Hopp T. P., Woods K. R. Prediction of protein antigenic determinants from amino acid sequences. Proc Natl Acad Sci U S A. 1981 Jun;78(6):3824–3828. doi: 10.1073/pnas.78.6.3824. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Kieff E. D., Bachenheimer S. L., Roizman B. Size, composition, and structure of the deoxyribonucleic acid of herpes simplex virus subtypes 1 and 2. J Virol. 1971 Aug;8(2):125–132. doi: 10.1128/jvi.8.2.125-132.1971. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Knipe D. M., Batterson W., Nosal C., Roizman B., Buchan A. Molecular genetics of herpes simplex virus. VI. Characterization of a temperature-sensitive mutant defective in the expression of all early viral gene products. J Virol. 1981 May;38(2):539–547. doi: 10.1128/jvi.38.2.539-547.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Knipe D. M., Ruyechan W. T., Honess R. W., Roizman B. Molecular genetics of herpes simplex virus: the terminal a sequences of the L and S components are obligatorily identical and constitute a part of a structural gene mapping predominantly in the S component. Proc Natl Acad Sci U S A. 1979 Sep;76(9):4534–4538. doi: 10.1073/pnas.76.9.4534. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Koment R. W., Rapp F. Temperature-sensitive host range mutants of herpes simplex virus type 2. J Virol. 1975 Apr;15(4):812–819. doi: 10.1128/jvi.15.4.812-819.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Liu F. T., Zinnecker M., Hamaoka T., Katz D. H. New procedures for preparation and isolation of conjugates of proteins and a synthetic copolymer of D-amino acids and immunochemical characterization of such conjugates. Biochemistry. 1979 Feb 20;18(4):690–693. doi: 10.1021/bi00571a022. [DOI] [PubMed] [Google Scholar]
  16. Mackem S., Roizman B. Regulation of herpesvirus macromolecular synthesis: temporal order of transcription of alpha genes is not dependent on the stringency of inhibition of protein synthesis. J Virol. 1981 Oct;40(1):319–322. doi: 10.1128/jvi.40.1.319-322.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Mackem S., Roizman B. Regulation of herpesvirus macromolecular synthesis: transcription-initiation sites and domains of alpha genes. Proc Natl Acad Sci U S A. 1980 Dec;77(12):7122–7126. doi: 10.1073/pnas.77.12.7122. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. McGeoch D. J., Dolan A., Donald S., Rixon F. J. Sequence determination and genetic content of the short unique region in the genome of herpes simplex virus type 1. J Mol Biol. 1985 Jan 5;181(1):1–13. doi: 10.1016/0022-2836(85)90320-1. [DOI] [PubMed] [Google Scholar]
  19. Morse L. S., Pereira L., Roizman B., Schaffer P. A. Anatomy of herpes simplex virus (HSV) DNA. X. Mapping of viral genes by analysis of polypeptides and functions specified by HSV-1 X HSV-2 recombinants. J Virol. 1978 May;26(2):389–410. doi: 10.1128/jvi.26.2.389-410.1978. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Murchie M. J., McGeoch D. J. DNA sequence analysis of an immediate-early gene region of the herpes simplex virus type 1 genome (map coordinates 0.950 to 0.978). J Gen Virol. 1982 Sep;62(Pt 1):1–15. doi: 10.1099/0022-1317-62-1-1. [DOI] [PubMed] [Google Scholar]
  21. Pellett P. E., Kousoulas K. G., Pereira L., Roizman B. Anatomy of the herpes simplex virus 1 strain F glycoprotein B gene: primary sequence and predicted protein structure of the wild type and of monoclonal antibody-resistant mutants. J Virol. 1985 Jan;53(1):243–253. doi: 10.1128/jvi.53.1.243-253.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Pereira L., Wolff M. H., Fenwick M., Roizman B. Regulation of herpesvirus macromolecular synthesis. V. Properties of alpha polypeptides made in HSV-1 and HSV-2 infected cells. Virology. 1977 Apr;77(2):733–749. doi: 10.1016/0042-6822(77)90495-0. [DOI] [PubMed] [Google Scholar]
  23. Post L. E., Mackem S., Roizman B. Regulation of alpha genes of herpes simplex virus: expression of chimeric genes produced by fusion of thymidine kinase with alpha gene promoters. Cell. 1981 May;24(2):555–565. doi: 10.1016/0092-8674(81)90346-9. [DOI] [PubMed] [Google Scholar]
  24. Post L. E., Roizman B. A generalized technique for deletion of specific genes in large genomes: alpha gene 22 of herpes simplex virus 1 is not essential for growth. Cell. 1981 Jul;25(1):227–232. doi: 10.1016/0092-8674(81)90247-6. [DOI] [PubMed] [Google Scholar]
  25. Preston C. M. Control of herpes simplex virus type 1 mRNA synthesis in cells infected with wild-type virus or the temperature-sensitive mutant tsK. J Virol. 1979 Jan;29(1):275–284. doi: 10.1128/jvi.29.1.275-284.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Sears A. E., Halliburton I. W., Meignier B., Silver S., Roizman B. Herpes simplex virus 1 mutant deleted in the alpha 22 gene: growth and gene expression in permissive and restrictive cells and establishment of latency in mice. J Virol. 1985 Aug;55(2):338–346. doi: 10.1128/jvi.55.2.338-346.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Spear P. G., Roizman B. Proteins specified by herpes simplex virus. V. Purification and structural proteins of the herpesvirion. J Virol. 1972 Jan;9(1):143–159. doi: 10.1128/jvi.9.1.143-159.1972. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Wilcox K. W., Kohn A., Sklyanskaya E., Roizman B. Herpes simplex virus phosphoproteins. I. Phosphate cycles on and off some viral polypeptides and can alter their affinity for DNA. J Virol. 1980 Jan;33(1):167–182. doi: 10.1128/jvi.33.1.167-182.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Journal of Virology are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES