title: Overexpression of active Syrian golden hamster prion protein PrPc as a glutathione S-transferase fusion in heterologous systems.
abstract: This article describes a procedure which permits for the first time the isolation of the prion protein PrPc from the Syrian golden hamster in heterologous systems. Using a glutathione S-transferase (GST) fusion approach, milligram amounts of stable, soluble, and homogeneous GST::PrPc protein were obtained in Escherichia coli and with baculovirus-infected insect cells. Authentic PrPc was released from the immobilized fusion protein by direct cleavage with thrombin. GST::PrPc expressed in these two expression systems and also authentic PrPc released by thrombin cleavage were recognized by a polyclonal antibody directed against amino acid 95 to 110 of the golden hamster PrPc protein. GST::PrPc was not detected by a monoclonal antibody recognizing the region encompassing amino acids 138 to 152 of the human prion protein. The fusion protein was sensitive to proteinase K digestion, demonstrating that the cellular rather than the proteinase K-resistant scrapie isoform was produced.
authors.affiliation: Laboratorium für Molekulare Biologie-Genzentrum-Institut für Biochemie der LMU München, Germany.
authors.forename: S
authors.initials: S
authors.lastname: Weiss
authors.forename: M
authors.initials: M
authors.lastname: Famulok
authors.forename: F
authors.initials: F
authors.lastname: Edenhofer
authors.forename: Y H
authors.initials: YH
authors.lastname: Wang
authors.forename: I M
authors.initials: IM
authors.lastname: Jones
authors.forename: M
authors.initials: M
authors.lastname: Groschup
authors.forename: E L
authors.initials: EL
authors.lastname: Winnacker
mesh_terms: D000818:Animals; D016367:Baculoviridae / Q000235:genetics; D002460:Cell Line; D003001:Cloning, Molecular; D006224:Cricetinae; D004926:Escherichia coli / Q000235:genetics; D005982:Glutathione Transferase / Q000235:genetics*; D008647:Mesocricetus; D010843:Pichia / Q000235:genetics; D011328:Prions / Q000235:genetics*; D011993:Recombinant Fusion Proteins / Q000235:genetics; D018411:Spodoptera
chemical_list: D011328:Prions; D011993:Recombinant Fusion Proteins; D005982:Glutathione Transferase