Abstract
Fragmentation of amyloid polymers by the chaperone Hsp104 allows them to propagate as prions in yeast. The factors which determine the frequency of fragmentation are unclear, though it is often presumed to depend on the physical strength of prion polymers. Proteins with long polyglutamine stretches represent a tractable model for revealing sequence elements required for polymer fragmentation in yeast, since they form poorly fragmented amyloids. Here we show that interspersion of polyglutamine stretches with various amino acid residues differentially affects the in vivo formation and fragmentation of the respective amyloids. Aromatic residues tyrosine, tryptophan and phenylalanine strongly stimulated polymer fragmentation, leading to the appearance of oligomers as small as dimers. Alanine, methionine, cysteine, serine, threonine and histidine also enhanced fragmentation, while charged residues, proline, glycine and leucine inhibited polymerization. Our data indicate that fragmentation frequency primarily depends on the recognition of fragmentation-promoting residues by Hsp104 and/or its co-chaperones, rather than on the physical stability of polymers. This suggests that differential exposure of such residues to chaperones defines prion variant-specific differences in polymer fragmentation efficiency.
Highlights
Some proteins can undergo non-covalent polymerization coupled with conformational rearrangement, resulting in the formation of amyloid fibrils with regular cross-b-sheet structure
PolyQX proteins and their ability to polymerize We created a set of multicopy plasmids encoding various poly(QQQXQ) domains, where X is any amino acid residue (Table 1), fused to Sup35 MC domains
These proteins are further referred to as nQX, where n is the length of the polyQX stretch
Summary
Some proteins can undergo non-covalent polymerization coupled with conformational rearrangement, resulting in the formation of amyloid fibrils with regular cross-b-sheet structure. Such fibrils tend to stick together, forming intra- or extracellular amyloid aggregates. Prions were found in fungi, mostly in the yeast Saccharomyces cerevisiae, where they define various heritable phenotypes. Among them, [PSI+] is probably the best studied This prion is related to the heritable polymerization of translation termination factor eRF3, called Sup, which reduces efficiency of translation termination resulting in a nonsense-suppressor phenotype. Its amino-terminal N domain (amino acid residues (aa) 1–123), called prion domain (PrD), is responsible for the prion properties of the protein, being necessary for its polymerization both in vivo [5] and in vitro [6]. The Sup PrD can be split into two areas, one of which (aa 1–40) is especially rich in glutamine (Q) and asparagine (N), while another
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