Peptide aptamers expressed in the secretory pathway interfere with cellular PrPSc formation.

Abstract

Prion diseases are rare and obligatory fatal neurodegenerative disorders caused by the accumulation of a misfolded isoform (PrPSc) of the host-encoded prion protein (PrPc). Prophylactic and therapeutic regimens against prion diseases are very limited. To extend such strategies we selected peptide aptamers binding to PrP from a combinatorial peptide library presented on the Escherichia coli thioredoxin A (trxA) protein as a scaffold. In a yeast two-hybrid screen employing full-length murine PrP (aa 23-231) as a bait we identified three peptide aptamers that reproducibly bind to PrP. Treatment of prion-infected cells with recombinantly expressed aptamers added to the culture medium abolished PrPSc conversion with an IC50 between 350 and 700 nM. For expression in eukaryotic cells, peptide aptamers were fused to an N-terminal signal peptide for entry of the secretory pathway. The C terminus was modified by a glycosyl-phosphatidyl-inositol-(GPI) anchoring signal, a KDEL retention motif and the transmembrane and cytosolic domain of LAMP-I, respectively. These peptide aptamers retained their binding properties to PrPc and, depending on peptide sequence and C-terminal modification, interfered with endogenous PrPSc conversion upon expression in prion-infected cells. Notably, infection of cell cultures could be prevented by expression of KDEL peptide aptamers. For the first time, we show that trxA-based peptide aptamers can be targeted to the secretory pathway, thereby not losing the affinity for their target protein. Beside their inhibitory effect on prion conversion, these molecules could be used as fundament for rational drug design.

• Authors: Sabine Gilch, Claudia Kehler, Hermann M Schätzl
• Journal: Journal of molecular biology (J Mol Biol) Vol. 371 Issue 2 Pg. 362-73 (Aug 10 2007) ISSN: 0022-2836 [Print] Netherlands
• PMID: 17574575 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• Aptamers, Peptide
• Glycosylphosphatidylinositols
• PrPSc Proteins
• Recombinant Proteins
• Thioredoxins

Topics

• Amino Acid Sequence
• Animals
• Aptamers, Peptide (chemistry, genetics, metabolism)
• Base Sequence
• Cell Line
• Cell Membrane (metabolism)
• Endoplasmic Reticulum (metabolism)
• Gene Expression Regulation
• Glycosylphosphatidylinositols (metabolism)
• Lysosomes (metabolism)
• Molecular Sequence Data
• PrPSc Proteins (genetics, metabolism)
• Protein Binding
• Rats
• Recombinant Proteins (genetics, metabolism)
• Thioredoxins (genetics, metabolism)