Comparative Analysis of the Human and Chicken Prion Protein Copper Binding Regions at pH 6.5

Lars Redecke,Wolfram Meyer-Klaucke,Mirjam Koker,Joachim Clos,Dessislava Georgieva,Nicolay Genov,Hartmut Echner,Hubert Kalbacher,Markus Perbandt,Reinhard Bredehorst,Wolfgang Voelter,Christian Betzel

doi:10.1074/jbc.m411775200

Abstract

Recent experimental evidence supports the hypothesis that prion proteins (PrPs) are involved in the Cu(II) metabolism. Moreover, the copper binding region has been implicated in transmissible spongiform encephalopathies, which are caused by the infectious isoform of prion proteins (PrP(Sc)). In contrast to mammalian PrP, avian prion proteins have a considerably different N-terminal copper binding region and, most interestingly, are not able to undergo the conversion process into an infectious isoform. Therefore, we applied x-ray absorption spectroscopy to analyze in detail the Cu(II) geometry of selected synthetic human PrP Cu(II) octapeptide complexes in comparison with the corresponding chicken PrP hexapeptide complexes at pH 6.5, which mimics the conditions in the endocytic compartments of neuronal cells. Our results revealed that structure and coordination of the human PrP copper binding sites are highly conserved in the pH 6.5-7.4 range, indicating that the reported pH dependence of copper binding to PrP becomes significant at lower pH values. Furthermore, the different chicken PrP hexarepeat motifs display homologous Cu(II) coordination at sub-stoichiometric copper concentrations. Regarding the fully cation-saturated prion proteins, however, a reduced copper coordination capability is supposed for the chicken prion protein based on the observation that chicken PrP is not able to form an intra-repeat Cu(II) binding site. These results provide new insights into the prion protein structure-function relationship and the conversion process of PrP.

Highlights

Recent experimental evidence supports the hypothesis that prion proteins (PrPs) are involved in the Cu(II) metabolism
We applied x-ray absorption spectroscopy to analyze in detail the Cu(II) geometry of selected synthetic human PrP Cu(II) octapeptide complexes in comparison with the corresponding chicken PrP hexapeptide complexes at pH 6.5, which mimics the conditions in the endocytic compartments of neuronal cells
Our results suggest that the structure of Cu(II) binding to the selected human octarepeat peptides differs only in the amount of involved histidine residues, depending on the number of repeat copies present within a single peptide. This is indicated by the extended x-ray absorption fine structure (EXAFS) data and by the qualitative analysis of the corresponding x-ray absorption near-end structure pattern

Summary

Introduction

Recent experimental evidence supports the hypothesis that prion proteins (PrPs) are involved in the Cu(II) metabolism. A total of five high affinity copper binding motifs of PrPC were identified, one at the beginning of the C-terminal domain centered at residues His-96 and His-111 [24, 25] and four within the His-containing sequence PHGGGWGQ that is repeated four times between residues 60 and 91 (9, 10, 26 –31) This N-terminal octapeptide repeat region, which is highly conserved in mammals [32, 33], cooperatively binds up to four Cu(II) ions with dissociation constants in the nanomolar to micromolar range, reflecting a significant pH-dependence [9, 26, 31, 34]. Morante et al [35] suggested that the coordination shell of Cu(II) varies depending on the occupancy of the available metal sites This x-ray absorption spectroscopy analysis of bovine octapeptide complexes linked the two common copper binding models, which differ mainly in the number of coordinated imidazole side chains

Methods

Results

Conclusion