Molecular Biotechnology

, Volume 60, Issue 8, pp 576–584 | Cite as

Effect of Temporal Expression of Integral Membrane Proteins by Baculovirus Expression Vector System

  • T. Z. SalemEmail author
  • F. Zhang
  • N. Sahly
  • S. Thiem
Original Paper


Integral membrane proteins (IMPs) are popular target for drugs, but their resolved structures have been overlooked when compared with cytosolic proteins. The main reason is that IMPs usually need intensive post-translational modifications and they are bound to membranes, which increase the complexity of purifying or crystalizing them. Although different expression systems are used to express IMPs, baculovirus is considered one of the most successful expression systems for those proteins. Despite that, there are always unknown discrepancies in the level of IMPs expression in the baculovirus expression system. Retrospective studies have shown that expression of an immunoglobulin (anti-Chymase mouse monoclonal IgG1) driven by vp39 promoter was more efficient compared to its expression under polyhedrin (polh) promoter; however, this conclusion was not tested on different IMPs to generalize such a conclusion. In this study, the expression of eight different IMPs has been compared under vp39 and polh promoters of Autographa californica nucleopolyhedrovirus. Although different IMPs have shown different patterns of expression, the expression driven by vp39 promoter was found to be generally more efficient than the polh promoter.


IMPs Vp39 promoter Polyhedrin promoter Baculovirus expression system Sf21 insect cells 


  1. 1.
    Bernaudat, F., Frelet-Barrand, A., Pochon, N., Dementin, S., Hivin, P., Boutigny, S., … Rolland, N. (2011). Heterologous expression of membrane proteins: Choosing the appropriate host. PLoS ONE, 6(12), e29191.CrossRefPubMedPubMedCentralGoogle Scholar
  2. 2.
    Griffith, D. A., Delipala, C., Leadsham, J., Jarvis, S. M., & Oesterhelt, D. (2003). A novel yeast expression system for the overproduction of quality-controlled membrane proteins. FEBS Letters, 553(1–2), 45–50.CrossRefPubMedGoogle Scholar
  3. 3.
    Sarramegna, V., Talmont, F., Demange, P., & Milon, A. (2003). Heterologous expression of G-protein-coupled receptors: Comparison of expression systems from the standpoint of large-scale production and purification. Cellular and Molecular Life Sciences (CMLS), 60(8), 1529–1546.CrossRefGoogle Scholar
  4. 4.
    Hopkins, R., Esposito, D., & Gillette, W. (2010). Widening the bottleneck: Increasing success in protein expression and purification. Journal of Structural Biology, 172(1), 14–20.CrossRefPubMedPubMedCentralGoogle Scholar
  5. 5.
    Quant, R. L., Pearson, M. N., Rohrmann, G. F., & Beaudreau, G. S. (1984). Production of polyhedrin monoclonal antibodies for distinguishing two Orgyia pseudotsugata baculoviruses. Applied and Environmental Microbiology, 48(4), 732–736.PubMedPubMedCentralGoogle Scholar
  6. 6.
    Summers, M. D., & Smith, G. E. (1987). A manual of methods for baculovirus vectors and insect cell culture procedures. College Station: Texas Agricultural and Mechanical College System.Google Scholar
  7. 7.
    Godeau, F., Casanova, J. L., Fairchild, K. D., Saucier, C., Delarbre, C., Gachelin, G., & Kourilsky, P. (1991). Expression and characterization of recombinant mouse beta 2-microglobulin type a in insect cells infected with recombinant baculoviruses. Research in Immunology, 142(5–6), 409–416.CrossRefPubMedGoogle Scholar
  8. 8.
    Higgins, M. K., Demir, M., & Tate, C. G. (2003). Calnexin co-expression and the use of weaker promoters increase the expression of correctly assembled Shaker potassium channel in insect cells. Biochimica et Biophysica Acta (BBA) - Biomembranes, 1610(1), 124–132.CrossRefGoogle Scholar
  9. 9.
    Jarvis, D. L., & Summers, M. D. (1989). Glycosylation and secretion of human tissue plasminogen activator in recombinant baculovirus-infected insect cells. Molecular and Cellular Biology, 9(1), 214–223.CrossRefPubMedPubMedCentralGoogle Scholar
  10. 10.
    Tate, C. G., Haase, J., Baker, C., Boorsma, M., Magnani, F., Vallis, Y., & Williams, D. C. (2003). Comparison of seven different heterologous protein expression systems for the production of the serotonin transporter. Biochimica et Biophysica Acta, 1610(1), 141–153.CrossRefPubMedGoogle Scholar
  11. 11.
    van Oers, M. M., Thomas, A. A., Moormann, R. J., & Vlak, J. M. (2001). Secretory pathway limits the enhanced expression of classical swine fever virus E2 glycoprotein in insect cells. Journal of Biotechnology, 86(1), 31–38.CrossRefPubMedGoogle Scholar
  12. 12.
    Grünewald, S., Haase, W., Reiländer, H., & Michel, H. (1996). Glycosylation, palmitoylation, and localization of the human D2S receptor in baculovirus-infected insect cells. Biochemistry, 35(48), 15149–15161.CrossRefPubMedGoogle Scholar
  13. 13.
    Ishiyama, S., & Ikeda, M. (2010). High-level expression and improved folding of proteins by using the vp39 late promoter enhanced with homologous DNA regions. Biotechnology Letters, 32(11), 1637–1647.CrossRefPubMedGoogle Scholar
  14. 14.
    Salem, T. Z., Zhang, F., & Thiem, S. M. (2013). Reduced expression of Autographa californica nucleopolyhedrovirus ORF34, an essential gene, enhances heterologous gene expression. Virology, 435(2), 225-238.CrossRefGoogle Scholar
  15. 15.
    O’Reilly, D. R., Miller, L., & Luckow, V. A. (1993). Baculovirus expression vectors: A laboratory manual. Oxford: Oxford University Press.Google Scholar
  16. 16.
    Drew, D., Lerch, M., Kunji, E., Slotboom, D.-J., & de Gier, J.-W. (2006). Optimization of membrane protein overexpression and purification using GFP fusions. Nature Methods, 3(4), 303–313.CrossRefPubMedGoogle Scholar
  17. 17.
    Xu, H., Fu, Y., Tian, W., & Cohen, D. M. (2006). Glycosylation of the osmoresponsive transient receptor potential channel TRPV4 on Asn-651 influences membrane trafficking. American Journal of Physiology-Renal Physiology, 290(5), F1103–F1109.CrossRefPubMedGoogle Scholar
  18. 18.
    Abrol, N., Smolin, N., Armanious, G., Ceholski, D. K., Trieber, C. A., Young, H. S., & Robia, S. L. (2014). Phospholamban C-terminal residues are critical determinants of the structure and function of the calcium ATPase regulatory complex. The Journal of Biological Chemistry, 289(37), 25855–25866.CrossRefPubMedPubMedCentralGoogle Scholar
  19. 19.
    Ying, J., Sharov, V., Xu, S., Jiang, B., Gerrity, R., Schöneich, C., & Cohen, R. A. (2008). Cysteine-674 oxidation and degradation of sarcoplasmic reticulum Ca2+ ATPase in diabetic pig aorta. Free Radical Biology and Medicine, 45(6), 756–762.CrossRefPubMedGoogle Scholar
  20. 20.
    Stults, J. T., O’Connell, K. L., Garcia, C., Wong, S., Engel, A. M., Garbers, D. L., & Lowe, D. G. (1994). The disulfide linkages and glycosylation sites of the human natriuretic peptide receptor-C homodimer. Biochemistry, 33(37), 11372–11381.CrossRefPubMedGoogle Scholar
  21. 21.
    Lesch, K. P., Wolozin, B. L., Murphy, D. L., & Reiderer, P. (1993). Primary structure of the human platelet serotonin uptake site: Identity with the brain serotonin transporter. Journal of Neurochemistry, 60(6), 2319–2322.CrossRefPubMedGoogle Scholar
  22. 22.
    Shim, J.-Y. (2009). Transmembrane helical domain of the cannabinoid CB1 receptor. Biophysical Journal, 96(8), 3251–3262.CrossRefPubMedPubMedCentralGoogle Scholar
  23. 23.
    Papoucheva, E., Dumuis, A., Sebben, M., Richter, D. W., & Ponimaskin, E. G. (2004). The 5-hydroxytryptamine(1A) receptor is stably palmitoylated, and acylation is critical for communication of receptor with Gi protein. The Journal of Biological Chemistry, 279(5), 3280–3291.CrossRefPubMedGoogle Scholar
  24. 24.
    Farzan, M., Mirzabekov, T., Kolchinsky, P., Wyatt, R., Cayabyab, M., Gerard, N. P., … Choe, H. (1999). Tyrosine sulfation of the amino terminus of CCR5 facilitates HIV-1 entry. Cell, 96(5), 667–676.CrossRefPubMedGoogle Scholar
  25. 25.
    Hereld, D., Vaughan, R., Kim, J. Y., Borleis, J., & Devreotes, P. (1994). Localization of ligand-induced phosphorylation sites to serine clusters in the C-terminal domain of the Dictyostelium cAMP receptor, cAR1. The Journal of Biological Chemistry, 269(9), 7036–7044.PubMedGoogle Scholar
  26. 26.
    Meusser, B., Hirsch, C., Jarosch, E., & Sommer, T. (2005). ERAD: The long road to destruction. Nature Cell Biology, 7(8), 766–772.CrossRefPubMedGoogle Scholar

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© Springer Science+Business Media, LLC, part of Springer Nature 2018

Authors and Affiliations

  1. 1.Biomedical SciencesUniversity of Science and Technology at Zewail CityGizaEgypt
  2. 2.Department of Microbial GeneticsAGERI, Agricultural Research CenterGizaEgypt
  3. 3.Department of EntomologyMichigan State UniversityEast LansingUSA
  4. 4.Department of Microbiology and Molecular GeneticsMichigan State UniversityEast LansingUSA

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