2.50
Hdl Handle:
http://hdl.handle.net/10033/297222
Title:
Multi-host expression system for recombinant production of challenging proteins.
Authors:
Meyer, Steffen; Lorenz, Carmen; Baser, Bahar; Wördehoff, Mona; Jäger, Volker; van den Heuvel, Joop
Abstract:
Recombinant production of complex eukaryotic proteins for structural analyses typically requires a profound screening process to identify suitable constructs for the expression of ample amounts of properly folded protein. Furthermore, the evaluation of an optimal expression host has a major impact on protein yield and quality as well as on actual cost of the production process. Here we present a novel fast expression system for multiple hosts based on a single donor vector termed pFlp-Bac-to-Mam. The range of applications of pFlp-Bac-to-Mam comprises highly efficient transient transfection of HEK293-6E in serum-free suspension culture and subsequent large-scale production of challenging proteins expressing in mg per Liter level using either the baculoviral expression vector system or stable CHO production cell lines generated by Flp-mediated cassette exchange. The success of the multi-host expression vector to identify the optimal expression strategy for efficient production of high quality protein is demonstrated in a comparative expression study of three model proteins representing different protein classes: intracellular expression using a fluorescent protein, secretion of a single-chain-Fv-hIgG1Fc fusion construct and production of a large amount of highly homogeneous protein sample of the extracellular domain of a Toll-like receptor. The evaluation of the production efficiency shows that the pFlp-Bac-to-Mam system allows a fast and individual optimization of the expression strategy for each protein class.
Affiliation:
Department of Molecular Structural Biology, Helmholtz Centre for Infection Research, Braunschweig, Germany.
Citation:
Multi-host expression system for recombinant production of challenging proteins. 2013, 8 (7):e68674 PLoS ONE
Journal:
PloS one
Issue Date:
2013
URI:
http://hdl.handle.net/10033/297222
DOI:
10.1371/journal.pone.0068674
PubMed ID:
23874717
Type:
Article
Language:
en
ISSN:
1932-6203
Appears in Collections:
Publications of the RG Rekombinante Proteinexpression (RPEX)

Full metadata record

DC FieldValue Language
dc.contributor.authorMeyer, Steffenen_GB
dc.contributor.authorLorenz, Carmenen_GB
dc.contributor.authorBaser, Baharen_GB
dc.contributor.authorWördehoff, Monaen_GB
dc.contributor.authorJäger, Volkeren_GB
dc.contributor.authorvan den Heuvel, Joopen_GB
dc.date.accessioned2013-08-01T10:49:35Z-
dc.date.available2013-08-01T10:49:35Z-
dc.date.issued2013-
dc.identifier.citationMulti-host expression system for recombinant production of challenging proteins. 2013, 8 (7):e68674 PLoS ONEen_GB
dc.identifier.issn1932-6203-
dc.identifier.pmid23874717-
dc.identifier.doi10.1371/journal.pone.0068674-
dc.identifier.urihttp://hdl.handle.net/10033/297222-
dc.description.abstractRecombinant production of complex eukaryotic proteins for structural analyses typically requires a profound screening process to identify suitable constructs for the expression of ample amounts of properly folded protein. Furthermore, the evaluation of an optimal expression host has a major impact on protein yield and quality as well as on actual cost of the production process. Here we present a novel fast expression system for multiple hosts based on a single donor vector termed pFlp-Bac-to-Mam. The range of applications of pFlp-Bac-to-Mam comprises highly efficient transient transfection of HEK293-6E in serum-free suspension culture and subsequent large-scale production of challenging proteins expressing in mg per Liter level using either the baculoviral expression vector system or stable CHO production cell lines generated by Flp-mediated cassette exchange. The success of the multi-host expression vector to identify the optimal expression strategy for efficient production of high quality protein is demonstrated in a comparative expression study of three model proteins representing different protein classes: intracellular expression using a fluorescent protein, secretion of a single-chain-Fv-hIgG1Fc fusion construct and production of a large amount of highly homogeneous protein sample of the extracellular domain of a Toll-like receptor. The evaluation of the production efficiency shows that the pFlp-Bac-to-Mam system allows a fast and individual optimization of the expression strategy for each protein class.en_GB
dc.language.isoenen
dc.rightsArchived with thanks to PloS oneen_GB
dc.titleMulti-host expression system for recombinant production of challenging proteins.en
dc.typeArticleen
dc.contributor.departmentDepartment of Molecular Structural Biology, Helmholtz Centre for Infection Research, Braunschweig, Germany.en_GB
dc.identifier.journalPloS oneen_GB

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