Baculovirus-driven protein expression in insect cells: A benchmarking study

  • J Struct Biol. 2018 Aug;203(2):71-80. doi: 10.1016/j.jsb.2018.03.004.
Peggy Stolt-Bergner  1 ,  Christian Benda  2 ,  Tim Bergbrede  3 ,  Hüseyin Besir  4 ,  Patrick H N Celie  5 ,  Cindy Chang  6 ,  David Drechsel  7 ,  Ariane Fischer  2 ,  Arie Geerlof  8 ,  Barbara Giabbai  9 ,  Joop van den Heuvel  10 ,  Georg Huber  8 ,  Wolfgang Knecht  11 ,  Anita Lehner  1 ,  Regis Lemaitre  7 ,  Kristina Nordén  11 ,  Gwynn Pardee  12 ,  Ines Racke  4 ,  Kim Remans  4 ,  Astrid Sander  13 ,  Judith Scholz  14 ,  Magda Stadnik  5 ,  Paola Storici  9 ,  Daniel Weinbruch  15 ,  Isabel Zaror  12 ,  Linda H L Lua  16 ,  Sabine Suppmann  17
Affiliations
  • 1. Vienna Biocenter Core Facilities GmbH, Dr. Bohrgasse 3, 1030 Vienna, Austria.
  • 2. Max-Planck Institute of Biochemistry, Structural Cell Biology Department, Am Klopferspitz 18, 82152 Martinsried, Germany.
  • 3. Lead Discovery Center GmbH, Otto-Hahn-Str. 15, 44227 Dortmund, Germany.
  • 4. European Molecular Biology Laboratory (EMBL) Heidelberg, Protein Expression and Purification Core Facility, 69117 Heidelberg, Germany.
  • 5. NKI Protein Facility, Division of Biochemistry, Netherlands Cancer Institute, Plesmanlaan 121, 1066 CX Amsterdam, The Netherlands.
  • 6. Protein Expression Facility, The University of Queensland, Brisbane, Queensland 4072, Australia.
  • 7. Protein Expression Purification and Characterization Facility, Max Planck Institute of Molecular Molecular Cell Biology and Genetics, Pfotenhauerstr. 108, D-01307 Dresden, Germany.
  • 8. Helmholtz Center Munich, Protein Expression and Purification Facility, Institute of Structural Biology, Ingolstädter Landstraße 1, D-85764 Neuherberg, Germany.
  • 9. Structural Biology Laboratory, Elettra-Sincrotrone Trieste S.C.p.A., Trieste 34149, Italy.
  • 10. Helmholtz Centre for Infection Research, Department Structure and Function of Proteins, Inhoffenstrasse 7, 38124 Braunschweig, Germany.
  • 11. Lund University, Lund Protein Production Platform (LP3), Sölvegatan 35, SE-223 62 Lund, Sweden.
  • 12. Novartis Institutes for BioMedical Research, Emeryville, CA, USA.
  • 13. Max Planck Institute of Molecular Physiology, Dortmund Protein Facility, Otto-Hahn-Str. 11, 44227 Dortmund, Germany.
  • 14. Max-Planck Institute of Biochemistry, Biochemistry Core Facility, Am Klopferspitz 18, 82152 Martinsried, Germany.
  • 15. Coriolis Pharma, Biopharmaceutical Research and Development Service, Am Klopferspitz 19, 82152 Martinsried, Germany.
  • 16. Protein Expression Facility, The University of Queensland, Brisbane, Queensland 4072, Australia. Electronic address: [email protected].
  • 17. Max-Planck Institute of Biochemistry, Biochemistry Core Facility, Am Klopferspitz 18, 82152 Martinsried, Germany. Electronic address: [email protected].
PMID: 29545204 DOI: 10.1016/j.jsb.2018.03.004
Abstract

Baculovirus-insect cell expression system has become one of the most widely used eukaryotic expression systems for heterologous protein production in many laboratories. The availability of robust insect cell lines, serum-free media, a range of vectors and commercially-packaged kits have supported the demand for maximizing the exploitation of the baculovirus-insect cell expression system. Naturally, this resulted in varied strategies adopted by different laboratories to optimize protein production. Most laboratories have preference in using either the E. coli transposition-based recombination bacmid technology (e.g. Bac-to-Bac®) or homologous recombination transfection within insect cells (e.g. flashBAC™). Limited data is presented in the literature to benchmark the protocols used for these baculovirus vectors to facilitate the selection of a system for optimal production of target proteins. Taking advantage of the Protein Production and Purification Partnership in Europe (P4EU) scientific network, a benchmarking initiative was designed to compare the diverse protocols established in thirteen individual laboratories. This benchmarking initiative compared the expression of four selected intracellular proteins (mouse Dicer-2, 204 kDa; human ABL1 wildtype, 126 kDa; human FMRP, 68 kDa; viral vNS1-H1, 76 kDa). Here, we present the expression and purification results on these proteins and highlight the significant differences in expression yields obtained using different commercially-packaged baculovirus vectors. The highest expression level for difficult-to-express intracellular protein candidates were observed with the EmBacY baculovirus vector system.

Keywords
Baculovirus-insect cell system; Benchmark.