Abstract
Methods for generating recombinant vaccinia viruses for the expression of foreign viral glycoproteins in mammalian cell lines and the purification of expressed viral glycoproteins are described. These methods are based on many years of experience with the influenza hemagglutinin glycoprotein (HA). However, they are applicable for studies on other viral glycoproteins, and with slight modifications, could be useful for cellular proteins as well.
This is a preview of subscription content, log in via an institution to check access.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Mackett, M., Smith, G. L., and Moss, B. (1982) Vaccinia virus: a selectable eukaryotic cloning and expression vector. Proc. Natl. Acad. Sci. USA 79, 7415–7419.
Moss, B. (1996) Genetically engineered poxviruses for recombinant gene expression, vaccination, and safety. Proc. Natl. Acad. Sci. USA 93, 11,341–11,348.
Blasco, R. and Moss, B. (1995) Selection of recombinant vaccinia viruses on the basis of plaque formation. Gene 158, 157–162.
Blasco, R. and Moss, B. (1991) Extracellular vaccinia virus formation and cell-to-cell virus transmission are prevented by deletion of the gene encoding the 37,000-Dalton outer envelope protein. J. Virol. 65, 5910–5920.
Cross, K. J., Wharton, S. A., Skehel, J. J., Wiley, D. C., and Steinhauer, D. A. (2001) Studies on influenza haemagglutinin fusion peptide mutants generated by reverse genetics. EMBO J. 20, 4432–4442.
Martin, J., Wharton, S. A., Lin, Y. P., et al. (1998) Studies of the binding properties of influenza hemagglutinin receptor-site mutants. Virology 241, 101–111.
Skehel, J. J. and Wiley, D. C. (2000) Receptor binding and membrane fusion in virus entry: the influenza hemagglutinin. Ann. Rev. Biochem. 69, 531–569.
Chen, J., Lee, K. H., Steinhauer, D. A., Stevens, D. J., Skehel, J. J., and Wiley, D. C. (1998) Structure of the hemagglutinin precursor cleavage site, a determinant of influenza pathogenicity and the origin of the labile conformation. Cell 95, 409–417.
Gamblin, S. J., Haire, L. F., Russell, R. J., et al. (2004) The structure and receptor binding properties of the 1918 influenza hemagglutinin. Science 303, 1838–1842.
Daniels, R. S., Douglas, A. R., Skehel, J. J., and Wiley, D. C. (1983) Analyses of the antigenicity of influenza haemagglutinin at the pH optimum for virus-mediated membrane fusion. J. Gen. Virol. 64, 1657–1662.
Godley, L., Pfeifer, J., Steinhauer, D., et al. (1992) Introduction of intersubunit disulfide bonds in the membrane-distal region of the influenza hemagglutinin abolishes membrane fusion activity. Cell 68, 635–645.
Author information
Authors and Affiliations
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2007 Humana Press Inc., Totowa, NJ
About this protocol
Cite this protocol
Li, ZN., Steinhauer, D.A. (2007). Expression and Purification of Viral Glycoproteins Using Recombinant Vaccinia Viruses for Functional and Structural Studies. In: Sugrue, R.J. (eds) Glycovirology Protocols. Methods in Molecular Biology, vol 379. Humana Press. https://doi.org/10.1007/978-1-59745-393-6_6
Download citation
DOI: https://doi.org/10.1007/978-1-59745-393-6_6
Publisher Name: Humana Press
Print ISBN: 978-1-58829-590-3
Online ISBN: 978-1-59745-393-6
eBook Packages: Springer Protocols