Preferential Cu2+ Coordination by His96 and His111 Induces β-Sheet Formation in the Unstructured Amyloidogenic Region of the Prion Protein

The prion protein (PrP) is a Cu 2+ binding cell surface glycoprotein that can misfold into a β-sheet-rich conformation to cause prion diseases. The majority of copper binding studies have concentrated on the octarepeat region of PrP. However, using a range of spectroscopic techniques, we show that copper binds preferentially to an unstructured region of PrP between residues 90 and 115, outside of the octarepeat domain. Comparison of recombinant PrP with PrP-(91–115) indicates that this prion fragment is a good model for Cu 2+ binding to the full-length protein. In contrast to previous reports we show that Cu 2+ binds to this region of PrP with a nanomolar dissociation constant. NMR and EPR spectroscopy indicate a square-planar or square-pyramidal Cu 2+ coordination utilizing histidine residues. Studies with PrP analogues show that the high affinity site requires both His 96 and His 111 as Cu 2+ ligands, rather than a complex centered on His 96 as has been previously suggested. Our circular dichroism studies indicate a loss of irregular structure on copper coordination with an increase in β-sheet conformation. It has been shown that this unstructured region, between residues 90 and 120, is vital for prion propagation and different strains of prion disease have been linked with copper binding. The role of Cu 2+ in prion misfolding and disease must now be re-evaluated in the light of these findings.