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Reference: Yu Y, et al. (2011) Flexibility of the Ure2 prion domain is important for amyloid fibril formation. Biochem J 434(1):143-151

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Abstract

Ure2, the protein determinant of the Saccharomyces cerevisiae prion [URE3], has a natively disordered N-terminal domain that is important for prion formation in vivo and amyloid formation in vitro; the globular C-domain has a glutathione transferase-like fold. Here, we swapped the position of the N- and C- regions, with or without an intervening peptide linker, to create the Ure2 variants CLN-Ure2 and CN-Ure2, respectively. The native structural content and stability of the variants were the same as wild-type Ure2, as indicated by enzymatic activity, far-UV circular dichroism and equilibrium denaturation. CLN-Ure2 was able to form amyloid-like fibrils, but with a significantly longer lag time than wild-type Ure2; and the two proteins were unable to cross-seed. Under the same conditions, CN-Ure2 showed limited ability to form fibrils, but this was improved after addition of 0.03 M GdmCl. As for wild-type Ure2, allosteric enzyme activity was observed in fibrils of CLN-Ure2 and CN-Ure2, consistent with retention of native-like dimeric structure of the C-domains within the fibrils. Proteolytically digested fibrils of CLN-Ure2 and CN-Ure2 showed the same residual fibril core morphology as wild-type Ure2. The results suggest that the position of the prion domain affects the ability of Ure2 to form fibrils primarily due to effects on its flexibility.

Reference Type
Journal Article
Authors
Yu Y, Wang HY, Bai M, Perrett S
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