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Baculovirus-driven protein expression in insect cells: A benchmarking study.
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.
Authors : Stolt-Bergner Peggy , Benda Christian , Bergbrede Tim , Besir Hüseyin , Celie Patrick H N , Chang Cindy , Drechsel David , Fischer Ariane , Geerlof Arie , Giabbai Barbara , van den Heuvel Joop , Huber Georg , Knecht Wolfgang , Lehner Anita , Lemaitre Regis , Nordén Kristina , Pardee Gwynn , Racke Ines , Remans Kim , Sander Astrid , Scholz Judith , Stadnik Magda , Storici Paola , Weinbruch Daniel , Zaror Isabel , Lua Linda H L , Suppmann Sabine ,
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Pathways :
WP1049: G Protein Signaling Pathways
WP1165: G Protein Signaling Pathways
WP1263: Mitochondrial Gene Expression
WP1301: Mitochondrial Gene Expression
WP1368: Mitochondrial Gene Expression
WP1371: G Protein Signaling Pathways
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WP1644: DNA replication
WP1650: Fluorobenzoate degradation
WP1654: gamma-Hexachlorocyclohexane degradation
WP1657: Glycerolipid metabolism
WP1659: Glycine, serine and threonine metabolism
WP1661: Glyoxylate and dicarboxylate metabolism
WP1663: Homologous recombination
WP1665: Limonene and pinene degradation
WP1672: Mismatch repair
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Bibliography :
[29545204] Baculovirus-driven protein expression in insect cells: A benchmarking study.
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