Skip to main content
Journal of Virology logoLink to Journal of Virology
. 1997 Aug;71(8):6061–6067. doi: 10.1128/jvi.71.8.6061-6067.1997

Posttranslational processing and identification of a neutralization domain of the GP4 protein encoded by ORF4 of Lelystad virus.

J J Meulenberg 1, A P van Nieuwstadt 1, A van Essen-Zandbergen 1, J P Langeveld 1
PMCID: PMC191865  PMID: 9223499

Abstract

GP4 is a minor structural glycoprotein encoded by ORF4 of Lelystad virus (LV). When it was immunoprecipitated from cell lysates and extracellular virus of CL2621 cells infected with LV, it was shown to have an apparent molecular mass of approximately 28 and 31 kDa, respectively. This difference in size occurred because its core N-glycans were modified to complex type N-glycans during the transport of the protein through the endoplasmic reticulum and Golgi compartment. A panel of 15 neutralizing monoclonal antibodies (MAbs) reacted with the native GP4 protein expressed by LV and the recombinant GP4 protein expressed in a Semliki Forest virus expression system. However, these MAbs did not react with the GP4 protein of U.S. isolate VR2332. To map the binding site of the MAbs, chimeric constructs composed of ORF4 of LV and VR2332 were generated. The reactivity of these constructs indicated that all the MAbs were directed against a region spanning amino acids 40 to 79 of the GP4 protein of LV. Six MAbs reacted with solid-phase synthetic dodecapeptides. The core of this site consists of amino acids 59 to 67 (SAAQEKISF). Comparison of the amino acid sequences of GP4 proteins from various European and North American isolates indicated that the neutralization domain spanning amino acids 40 to 79 is the most variable region of GP4. The neutralization domain of GP4, described here, is the first identified for LV.

Full Text

The Full Text of this article is available as a PDF (660.4 KB).

Selected References

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

  1. Balasuriya U. B., Maclachlan N. J., De Vries A. A., Rossitto P. V., Rottier P. J. Identification of a neutralization site in the major envelope glycoprotein (GL) of equine arteritis virus. Virology. 1995 Mar 10;207(2):518–527. doi: 10.1006/viro.1995.1112. [DOI] [PubMed] [Google Scholar]
  2. Bautista E. M., Meulenberg J. J., Choi C. S., Molitor T. W. Structural polypeptides of the American (VR-2332) strain of porcine reproductive and respiratory syndrome virus. Arch Virol. 1996;141(7):1357–1365. doi: 10.1007/BF01718837. [DOI] [PubMed] [Google Scholar]
  3. Benfield D. A., Nelson E., Collins J. E., Harris L., Goyal S. M., Robison D., Christianson W. T., Morrison R. B., Gorcyca D., Chladek D. Characterization of swine infertility and respiratory syndrome (SIRS) virus (isolate ATCC VR-2332). J Vet Diagn Invest. 1992 Apr;4(2):127–133. doi: 10.1177/104063879200400202. [DOI] [PubMed] [Google Scholar]
  4. Collins J. E., Benfield D. A., Christianson W. T., Harris L., Hennings J. C., Shaw D. P., Goyal S. M., McCullough S., Morrison R. B., Joo H. S. Isolation of swine infertility and respiratory syndrome virus (isolate ATCC VR-2332) in North America and experimental reproduction of the disease in gnotobiotic pigs. J Vet Diagn Invest. 1992 Apr;4(2):117–126. doi: 10.1177/104063879200400201. [DOI] [PubMed] [Google Scholar]
  5. Conzelmann K. K., Visser N., Van Woensel P., Thiel H. J. Molecular characterization of porcine reproductive and respiratory syndrome virus, a member of the arterivirus group. Virology. 1993 Mar;193(1):329–339. doi: 10.1006/viro.1993.1129. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Deregt D., de Vries A. A., Raamsman M. J., Elmgren L. D., Rottier P. J. Monoclonal antibodies to equine arteritis virus proteins identify the GL protein as a target for virus neutralization. J Gen Virol. 1994 Sep;75(Pt 9):2439–2444. doi: 10.1099/0022-1317-75-9-2439. [DOI] [PubMed] [Google Scholar]
  7. Faaberg K. S., Even C., Palmer G. A., Plagemann P. G. Disulfide bonds between two envelope proteins of lactate dehydrogenase-elevating virus are essential for viral infectivity. J Virol. 1995 Jan;69(1):613–617. doi: 10.1128/jvi.69.1.613-617.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Faaberg K. S., Plagemann P. G. The envelope proteins of lactate dehydrogenase-elevating virus and their membrane topography. Virology. 1995 Oct 1;212(2):512–525. doi: 10.1006/viro.1995.1509. [DOI] [PubMed] [Google Scholar]
  9. Geysen H. M., Meloen R. H., Barteling S. J. Use of peptide synthesis to probe viral antigens for epitopes to a resolution of a single amino acid. Proc Natl Acad Sci U S A. 1984 Jul;81(13):3998–4002. doi: 10.1073/pnas.81.13.3998. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Glaser A. L., de Vries A. A., Dubovi E. J. Comparison of equine arteritis virus isolates using neutralizing monoclonal antibodies and identification of sequence changes in GL associated with neutralization resistance. J Gen Virol. 1995 Sep;76(Pt 9):2223–2233. doi: 10.1099/0022-1317-76-9-2223. [DOI] [PubMed] [Google Scholar]
  11. Harty J. T., Plagemann P. G. Formalin inactivation of the lactate dehydrogenase-elevating virus reveals a major neutralizing epitope not recognized during natural infection. J Virol. 1988 Sep;62(9):3210–3216. doi: 10.1128/jvi.62.9.3210-3216.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Hulst M. M., Westra D. F., Wensvoort G., Moormann R. J. Glycoprotein E1 of hog cholera virus expressed in insect cells protects swine from hog cholera. J Virol. 1993 Sep;67(9):5435–5442. doi: 10.1128/jvi.67.9.5435-5442.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Kim H. S., Kwang J., Yoon I. J., Joo H. S., Frey M. L. Enhanced replication of porcine reproductive and respiratory syndrome (PRRS) virus in a homogeneous subpopulation of MA-104 cell line. Arch Virol. 1993;133(3-4):477–483. doi: 10.1007/BF01313785. [DOI] [PubMed] [Google Scholar]
  14. Kuo L. L., Harty J. T., Erickson L., Palmer G. A., Plagemann P. G. A nested set of eight RNAs is formed in macrophages infected with lactate dehydrogenase-elevating virus. J Virol. 1991 Sep;65(9):5118–5123. doi: 10.1128/jvi.65.9.5118-5123.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  16. Langedijk J. P., Back N. K., Kinney-Thomas E., Bruck C., Francotte M., Goudsmit J., Meloen R. H. Comparison and fine mapping of both high and low neutralizing monoclonal antibodies against the principal neutralization domain of HIV-1. Arch Virol. 1992;126(1-4):129–146. doi: 10.1007/BF01309690. [DOI] [PubMed] [Google Scholar]
  17. Langeveld J. P., Casal J. I., Vela C., Dalsgaard K., Smale S. H., Puijk W. C., Meloen R. H. B-cell epitopes of canine parvovirus: distribution on the primary structure and exposure on the viral surface. J Virol. 1993 Feb;67(2):765–772. doi: 10.1128/jvi.67.2.765-772.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Liljeström P., Garoff H. A new generation of animal cell expression vectors based on the Semliki Forest virus replicon. Biotechnology (N Y) 1991 Dec;9(12):1356–1361. doi: 10.1038/nbt1291-1356. [DOI] [PubMed] [Google Scholar]
  19. Mardassi H., Massie B., Dea S. Intracellular synthesis, processing, and transport of proteins encoded by ORFs 5 to 7 of porcine reproductive and respiratory syndrome virus. Virology. 1996 Jul 1;221(1):98–112. doi: 10.1006/viro.1996.0356. [DOI] [PubMed] [Google Scholar]
  20. Meulenberg J. J., Petersen-den Besten A., De Kluyver E. P., Moormann R. J., Schaaper W. M., Wensvoort G. Characterization of proteins encoded by ORFs 2 to 7 of Lelystad virus. Virology. 1995 Jan 10;206(1):155–163. doi: 10.1016/S0042-6822(95)80030-1. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Meulenberg J. J., Petersen-den Besten A. Identification and characterization of a sixth structural protein of Lelystad virus: the glycoprotein GP2 encoded by ORF2 is incorporated in virus particles. Virology. 1996 Nov 1;225(1):44–51. doi: 10.1006/viro.1996.0573. [DOI] [PubMed] [Google Scholar]
  22. Meulenberg J. J., de Meijer E. J., Moormann R. J. Subgenomic RNAs of Lelystad virus contain a conserved leader-body junction sequence. J Gen Virol. 1993 Aug;74(Pt 8):1697–1701. doi: 10.1099/0022-1317-74-8-1697. [DOI] [PubMed] [Google Scholar]
  23. Murtaugh M. P., Elam M. R., Kakach L. T. Comparison of the structural protein coding sequences of the VR-2332 and Lelystad virus strains of the PRRS virus. Arch Virol. 1995;140(8):1451–1460. doi: 10.1007/BF01322671. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Nelson E. A., Christopher-Hennings J., Drew T., Wensvoort G., Collins J. E., Benfield D. A. Differentiation of U.S. and European isolates of porcine reproductive and respiratory syndrome virus by monoclonal antibodies. J Clin Microbiol. 1993 Dec;31(12):3184–3189. doi: 10.1128/jcm.31.12.3184-3189.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Plagemann P. G., Moennig V. Lactate dehydrogenase-elevating virus, equine arteritis virus, and simian hemorrhagic fever virus: a new group of positive-strand RNA viruses. Adv Virus Res. 1992;41:99–192. doi: 10.1016/S0065-3527(08)60036-6. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Wensvoort G., Terpstra C., Boonstra J., Bloemraad M., Van Zaane D. Production of monoclonal antibodies against swine fever virus and their use in laboratory diagnosis. Vet Microbiol. 1986 Jul;12(2):101–108. doi: 10.1016/0378-1135(86)90072-6. [DOI] [PubMed] [Google Scholar]
  27. Zeng L., Godeny E. K., Methven S. L., Brinton M. A. Analysis of simian hemorrhagic fever virus (SHFV) subgenomic RNAs, junction sequences, and 5' leader. Virology. 1995 Mar 10;207(2):543–548. doi: 10.1006/viro.1995.1114. [DOI] [PubMed] [Google Scholar]
  28. de Vries A. A., Chirnside E. D., Horzinek M. C., Rottier P. J. Structural proteins of equine arteritis virus. J Virol. 1992 Nov;66(11):6294–6303. doi: 10.1128/jvi.66.11.6294-6303.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. den Boon J. A., Snijder E. J., Chirnside E. D., de Vries A. A., Horzinek M. C., Spaan W. J. Equine arteritis virus is not a togavirus but belongs to the coronaviruslike superfamily. J Virol. 1991 Jun;65(6):2910–2920. doi: 10.1128/jvi.65.6.2910-2920.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. van Nieuwstadt A. P., Meulenberg J. J., van Essen-Zanbergen A., Petersen-den Besten A., Bende R. J., Moormann R. J., Wensvoort G. Proteins encoded by open reading frames 3 and 4 of the genome of Lelystad virus (Arteriviridae) are structural proteins of the virion. J Virol. 1996 Jul;70(7):4767–4772. doi: 10.1128/jvi.70.7.4767-4772.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]

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

RESOURCES