Overview of the Baculovirus Expression System

Cheryl Isaac Murphy1, Helen Piwnica‐Worms2, Stefan Grünwald3, William G. Romanow4, Nicole Francis5, Hua‐Ying Fan5

1 Aquila Biopharmaceuticals, Worcester, Massachusetts, 2 Washington University School of Medicine, St. Louis, Missouri, 3 Pharmingen, San Diego, California, 4 Corning Costar, Portsmouth, New Hampshire, 5 Massachusetts General Hospital, Boston, Massachusetts
Publication Name:  Current Protocols in Molecular Biology
Unit Number:  Unit 16.9
DOI:  10.1002/0471142727.mb1609s65
Online Posting Date:  February, 2004
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Abstract

Baculoviruses have emerged as a popular system for overproducing recombinant proteins in eukaryotic cells. This unit gives an overview of the baculovirus expression system, including discussion of the baculovirus life cycle, and postā€translational modifications that occur in insect cells. In addition, the steps for overproducing proteins in the baculovirus systems are described along with recommendations for choosing an appropriate baculovirus vector and DNA, and reagents and equipment necessary for implementing the whole overexpression system.

     
 
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Table of Contents

  • Expression of Proteins in Insect Cells Using Baculovirus Vectors
  • Baculovirus Life Cycle
  • Baculovirus Expression System
  • Posttranslational Modification of Proteins in Insect Cells
  • Steps for Overproducing Proteins Using the Baculovirus System
  • Choosing a Baculovirus Transfer Vector
  • Choosing a Baculovirus DNA
  • Reagents, Solutions, and Equipment for the Baculovirus System
  • Literature Cited
  • Figures
  • Tables
     
 
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Materials

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Figures

Videos

Literature Cited

   Carbonell, L.F., Klowden, M.J., and Miller, L.K. 1985. Baculovirus‐mediated expression of bacterial genes in dipteran and mammalian cells. J. Virol. 56:153‐160.
   Davidson, D.J.C., Fraser, M.J., and Castellino, F.J. 1990. Oligosaccharide processing in the expression of human plasminogen cDNA by lepidopteran insect (Spodoptera frugiperda) cells. Biochemistry 29:5584‐5590.
   Davis, T.R. and Wood, H.A. 1995. Intrinsic glycosylation potentials of insect‐cell cultures and insect larvae. In Vitro Cell. Dev. Biol. Animal 31:659‐663.
   Doerfler, W. and Bohm, P. 1986. The Molecular Biology of Baculoviruses. Springer‐Verlag, New York.
   Grabenhorst, E., Hofer, B., Nimitz, M., Jager, V., and Contradt, H.S. 1993. Biosynthesis and secretion of human interleukin‐2 glycoprotein variants from baculovirus‐infected insect Sf21 cells: Characterization of polypeptides and post‐translational modifications. Eur. J. Biochem. 215:189‐197.
   Hellers, M., Gunne, H., and Steiner, H. 1991. Expression and post‐translational processing of preprocecropin‐A using a baculovirus vector. Eur. J. Biochem. 199:435‐439.
   James, D.C., Freedman, R.B., Hoare, M., Ogonah, O.W., Rooney, B.C., and Larionov, O.A. 1995. N‐glycosylation of recombinant human interferon‐g produced in different animal expression systems. Bio/Technology 13:592‐596.
   Jarvis, D.L. and Finn, E.E. 1995. Biochemical analysis of the N‐glycosylation pathway in baculovirus‐infected lepidopteran insect cells. Virology 212:500‐511.
   Jarvis, D.L. and Summers, M.D. 1992. Baculovirus expression vectors. In Recombinant DNA Vaccines: Rationale and Strategies (R.E. Isaacson, ed.) pp. 265‐291. Marcel Dekker, New York.
   Kitts, P.A. and Possee, R.D. 1993. A method for producing recombinant baculovirus expression vectors at high frequency. BioTechniques 14:810‐817.
   Livingston, C. and Jones, I. 1989. Baculovirus expression vector with single‐strand capability. Nucl. Acids Res. 17:2366.
   Luckow, V.A. 1991. Cloning and expression of heterologous genes in insect cells with baculovirus vectors. In Recombinant DNA Technology and Applications (A. Prokop, R.K. Bajpai,, and, C. Ho, eds.) pp. 97‐152. McGraw‐Hill, New York.
   Luckow, V.A. and Summers, M.D. 1988. Trends in the development of baculoviral expression vectors. Bio/Technology 6:47‐55.
   Matsuura, Y., Possee, R.D., and Bishop, D.H.L. 1987. Baculovirus expression vectors: The requirements for high level expression of proteins, including glycoproteins. J. Gen. Virol. 68:1233‐1250.
   Miller, L.K. 1988. Baculoviruses as gene expression vectors. Annu. Rev. Microbiol. 42:177‐199.
   Miller, D.W., Safer, P., and Miller, L.K. 1986. An insect baculovirus host vector for high‐level expression of foreign genes. In Genetic Engineering, Vol. 8 (J.K. Setlow, and, A. Hollaender, eds.) pp. 277‐298. Plenum, New York.
   Murphy, C.I., McIntire, J.R., Davis, D.R., Hodgdon, H., Seals, J.R., and Young, E. 1993. Enhanced expression, secretion, and large‐scale purification of recombinant HIV‐1 gp120 in insect cells using the baculovirus egt and p67 signal peptides. Protein Expr. Purif. 4:349‐357.
   Ogonah, O.W., Freedman, R.B., Jenkins, N., Patel, K., and Rooney, B.C. 1996. Isolation and characterization of an insect cell line able to perform complex N‐linked glycosylation on recombinant proteins. Bio/Technology 14:197‐202.
   O'Reilly, D.R., Miller, L.K., and Luckow, V.A. 1992. Baculovirus Expression Vectors. W.H. Freeman, New York.
   Piwnica‐Worms, H., Williams, N.G., Cheng, S.H., and Roberts, T.M. 1990. Regulation of pp60 c‐src and its association with polyoma virus middle T antigen in insect cells. J. Virol. 64:61‐68.
   Summers, M.D. and Smith, G.E. 1987. A manual of methods for baculovirus vectors and insect cell culture procedures. Texas Agricultural Experiment Station Bulletin No.1555. College Station, Texas.
   Vialard, J., Lalumiere, M., Vernet, T., Briedis, D., Alkhatib, G., Henning, D., Levin, D., and Richardson, C. 1990. Synthesis of the membrane fusion and hemagglutinin proteins of measle virus, using a novel baculovirus vector containing the β‐galactosidase gene. J. Virol. 64:37‐50.
Key Reference
   O'Reilly et al., 1992. See above.
  A guide assembled to aid researchers using the baculoviral expression system, containing detailed protocols for using this system effectively.
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