title: Effect of charged residues in the N-domain of Sup35 protein on prion [PSI+] stability and propagation.
abstract: Recent studies have shown that Sup35p prion fibrils probably have a parallel in-register β-structure. However, the part(s) of the N-domain critical for fibril formation and maintenance of the [PSI(+)] phenotype remains unclear. Here we designed a set of five SUP35 mutant alleles (sup35(KK)) with lysine substitutions in each of five N-domain repeats, and investigated their effect on infectivity and ability of corresponding proteins to aggregate and coaggregate with wild type Sup35p in the [PSI(+)] strain. Alleles sup35-M1 (Y46K/Q47K) and sup35-M2 (Q61K/Q62K) led to prion loss, whereas sup35-M3 (Q70K/Q71K), sup35-M4 (Q80K/Q81K), and sup35-M5 (Q89K/Q90K) were able to maintain the [PSI(+)] prion. This suggests that the critical part of the parallel in-register β-structure for the studied [PSI(+)] prion variant lies in the first 63-69 residues. Our study also reveals an unexpected interplay between the wild type Sup35p and proteins expressed from the sup35(KK) alleles during prionization. Both Sup35-M1p and Sup35-M2p coaggregated with Sup35p, but only sup35-M2 led to prion loss in a dominant manner. We suggest that in the fibrils, Sup35p can bind to Sup35-M1p in the same conformation, whereas Sup35-M2p only allowed the Sup35p conformation that leads to the non-heritable fold. Mutations sup35-M4 and sup35-M5 influence the structure of the prion forming region to a lesser extent, and can lead to the formation of new prion variants.
authors: From the Department of Genetics and Biotechnology, St. Petersburg State University, Saint Petersburg, Russia.
authors: Stanislav A
authors: SA
authors: Bondarev
authors: Vadim V
authors: VV
authors: Shchepachev
authors: Andrey V
authors: AV
authors: Kajava
authors: Galina A
authors: GA
authors: Zhouravleva
mesh_terms: D000483:Alleles; D000595:Amino Acid Sequence; D000596:Amino Acids / Q000378:metabolism*; D006720:Homozygote; D007700:Kinetics; D008954:Models, Biological; D008969:Molecular Sequence Data; D050505:Mutant Proteins / Q000378:metabolism; D009154:Mutation / Q000235:genetics; D010454:Peptide Termination Factors / Q000737:chemistry* / Q000378:metabolism*; D010641:Phenotype; D011328:Prions / Q000737:chemistry / Q000378:metabolism*; D011485:Protein Binding; D055550:Protein Stability; D020836:Protein Structure, Quaternary; D017434:Protein Structure, Tertiary; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry* / Q000378:metabolism*; D013329:Structure-Activity Relationship
chemical_list: D000596:Amino Acids; D050505:Mutant Proteins; D010454:Peptide Termination Factors; D011328:Prions; C068068:SUP35 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins
keywords: Amyloid; Prions; Protein Misfolding; Protein Structure; Saccharomyces cerevisiae; Superpleated β-Structure; Translation Release Factors; Translation Termination; [PSI+]
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