UCLA

In yeast, the formation of Ure2 fibrils underlies the prion state [URE3], in which the yeast loses the ability to distinguish good nitrogen sources from bad ones. The Ure2 prion domain is both necessary and sufficient for the formation of amyloid fibrils. Understanding the structure of Ure2 fibrils is important for understanding the propagation not only of the [URE3] prion but also of other yeast prions whose prion domains share similar features, such as the enrichment of asparagine and glutamine residues. Here, we report a structural study of the amyloid fibrils formed by the Ure2 prion domain using site-directed spin labeling and electron paramagnetic resonance (EPR) spectroscopy. We completed a spin label scanning of all the residue positions between 2 and 80 of the Ure2 prion domain. The EPR data show that the Ure2 fibril core consists of residues 8-68 and adopts a parallel in-register β-sheet structure. Most of the residues show strong spin-exchange interactions, suggesting that there are only short turns and no long loops in the fibril core. Based on the strength of spin-exchange interactions, we determined the likely locations of the β-strands. EPR data also show that the C-terminal region of the Ure2 prion domain is more disordered than the N-terminal region. The roles of hydrophobic and charged residues are analyzed. Overall, the structure of Ure2 fibrils appears to involve a balance of stabilizing interactions, such as asparagine ladders, and destabilizing interactions, such as stacking of charged residues.