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J. Virol., 08 1995, 4776-4783, Vol 69, No. 8
Copyright © 1995, American Society for Microbiology

Overexpression of active Syrian golden hamster prion protein PrPc as a glutathione S-transferase fusion in heterologous systems

S Weiss, M Famulok, F Edenhofer, YH Wang, IM Jones, M Groschup and EL Winnacker
Laboratorium fur Molekulare Biologie-Genzentrum-Institut fur Biochemie der LMU Munchen, Germany.

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.

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