Abstract
The genome region encoding the major envelope glycoprotein E2 (gp55) of the classical swine fever virus (CSFV) strain Alfort/187 was cloned and sequenced. The E2 gene, either with or without additional authentic 5′-terminal sequences coding for two variants of a putative signal sequence, was used to construct recombinant baculoviruses expressing the respective glycosylated and nonglycosylated E2 protein in insect cells. The signal sequences mediated glycosylation in insect cells, but no efficient secretion of the protein into the cell culture supernatant was observed. Six histidine residues introduced at the carboxy terminus of E2 allowed purification of E2 protein by Ni2+-chelate affinity chromatography. The proteins obtained were characterized and their immunological properties were compared by western blot analysis.
Similar content being viewed by others
References
Francki R.I.B., Fauquet C.M., Knudson D.L., and Brown F., Arch Virol2(Suppl), 228–229, 1991.
Dahle J., Liess B., and Frey H.R. in Harkness J.W. (ed).Pestivirus Infections of Ruminants. Comm Eur Communities [Rep.] EUR, Luxembourg, EUR-10238 EN, 1987, pp. 195–211.
Phillip J.I.H. and Darbyshire J.H., J Comp Pathol82 105–109, 1972.
Snowdon W.A. and French E.L., Aust Vet J44 179–184, 1968.
Stewart W.C., Carbrey E.A., Jenny E.W., Brown C.L., and Kresse J.I., J Am Vet Med Assoc159 1556–1563, 1971.
Terpstra C. and Wensvoort G., Res Vet Sci45 137–142, 1988.
Snowdon W.A. and French E.L. in van Bekkum J.G. (ed).Diagnosis and Epizootiology of Classical Swine Fever. EEC, Luxembourg, EUR 5486, 1976, pp. 159–183. Seminar, Amsterdam, 30 April to 2 May 1975.
Moennig V. and Plagemann P.G.W., Adv Virus Res41 53–98, 1992.
Meyers G., Rümenapf T., and Thiel H.J., Virology171 555–567, 1989.
Moormann R.J.M., Warmerdam P.A.M., van der Meer B., Schaaper W.M.M., Wensvoort G., and Hulst M.M., Virology177 184–198, 1990.
Muyldermans G., San Gabriel M.C., Caij A., De Smet A., and Hamers R., Arch Virol132 429–435, 1993.
Greiser-Wilke I., Moennig V., Coulibaly C.O.Z., Dahle J., Leder L., and Liess B., Arch Virol111 213–225, 1990.
Aynaud J.M., Ann Rech Vet1 25–36, 1968.
Rümenapf T., Unger G., Strauss J.H., and Thiel H.J., J Virol67 3288–3294, 1993.
Wensvoort G., Boonstra J., and Bodzinga B.G., J Gen Virol17 531–540, 1990.
Hulst M.M., Westra D.F., Wensvoort G., and Moormann R.J.M., J Virol67 5435–5442, 1993.
von Heijne G., J Mol Biol184 99–105, 1985.
von Heijne G., Nucleic Acids Res14 4683–4690, 1986.
Kitts P.A. and Possee R.D., Bio Techniques14 810–817, 1993.
Summers M.D. and Smith G.E.,A Manual of Methods for Baculovirus Vectors and Insect Cell Culture Procedures. Texas Agricultural Experiment Station Bulletin No. 1555, College Station, TX, 1987.
Bolin S., Moennig V., Kelso Gourley N.E., and Ridpath J., Arch Virol99 117–123, 1988.
Wensvoort G., J Gen Virol70 2865–2876, 1989.
Kozak M., Cell44 283–292, 1986.
Saiki R.K., Scharf S., Faloona F., Mullis K.B., Horn G.T., Ehrlich H.A., and Arnheim N., Science230 1350–1354, 1985.
Sambrook J., Fritsch E.F., and Maniatis T.,Molecular Cloning: A Laboratory Manual 2nd ed. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, 1989.
Mitchell D.B., Ruggli N., and Tratschin J.D., PCR Methods Appl2 81–82, 1992.
Sanger F., Nicklen S., and Coulson A.R., Proc Natl Acad Sci USA74 5463–5467, 1977.
Webb N.R. and Summers M.D., Technique-A J Methods Cell Mol Biol2 173–188, 1990.
Ausubel F.M., Brent R., Kingston R.E., Moore D.D., Seidmann J.G., Smith J.A., and Struhl K. (eds).Current Protocols in Molecular Biology John Wiley, New York, 1987.
Hochuli E., Bannwarth W., Döbeli H., Gentz R., and Stüber D., Bio/Technology6 1321–1325, 1988.
Have P., Acta Vet Scand25 462–465, 1984.
Wensvoort G., Bloemraad M., and Terpstra C., Vet Microbiol17 129–140, 1988.
Hofmann M.A., Brechtbühl K., and Stäuber N., Arch Virol139 217–229, 1994.
Flamand M., Deubel V., and Girard M., Virology191 826–836, 1992.
Lodisch H.F., J Biol Chem263 2107–2110, 1988.
Desprès Ph., Girard M., and Bouloy M., J Gen Virol72 1331–1342, 1991.
Mason P.W., Virology169 354–364, 1989.
Author information
Authors and Affiliations
Corresponding author
Rights and permissions
About this article
Cite this article
Ruggli, N., Moser, C., Mitchell, D. et al. Baculovirus expression and affinity purification of protein E2 of classical swine fever virus strain Alfort/187. Virus Genes 10, 115–126 (1995). https://doi.org/10.1007/BF01702592
Received:
Accepted:
Issue Date:
DOI: https://doi.org/10.1007/BF01702592