CUED Publications database

Influence of specific HSP70 domains on fibril formation of the yeast prion protein Ure2.

Xu, LQ and Wu, S and Buell, AK and Cohen, SI and Chen, LJ and Hu, WH and Cusack, SA and Itzhaki, LS and Zhang, H and Knowles, TP and Dobson, CM and Welland, ME and Jones, GW and Perrett, S (2013) Influence of specific HSP70 domains on fibril formation of the yeast prion protein Ure2. Philos Trans R Soc Lond B Biol Sci, 368. 20110410-.

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Abstract

Ure2p is the protein determinant of the Saccharomyces cerevisiae prion state [URE3]. Constitutive overexpression of the HSP70 family member SSA1 cures cells of [URE3]. Here, we show that Ssa1p increases the lag time of Ure2p fibril formation in vitro in the presence or absence of nucleotide. The presence of the HSP40 co-chaperone Ydj1p has an additive effect on the inhibition of Ure2p fibril formation, whereas the Ydj1p H34Q mutant shows reduced inhibition alone and in combination with Ssa1p. In order to investigate the structural basis of these effects, we constructed and tested an Ssa1p mutant lacking the ATPase domain, as well as a series of C-terminal truncation mutants. The results indicate that Ssa1p can bind to Ure2p and delay fibril formation even in the absence of the ATPase domain, but interaction of Ure2p with the substrate-binding domain is strongly influenced by the C-terminal lid region. Dynamic light scattering, quartz crystal microbalance assays, pull-down assays and kinetic analysis indicate that Ssa1p interacts with both native Ure2p and fibril seeds, and reduces the rate of Ure2p fibril elongation in a concentration-dependent manner. These results provide new insights into the structural and mechanistic basis for inhibition of Ure2p fibril formation by Ssa1p and Ydj1p.

Item Type: Article
Uncontrolled Keywords: Adenosine Triphosphatases Biosensing Techniques Gene Expression Regulation, Fungal Glutathione Peroxidase HSP70 Heat-Shock Proteins Kinetics Models, Molecular Molecular Chaperones Prions Protein Conformation Protein Structure, Tertiary Saccharomyces cerevisiae Saccharomyces cerevisiae Proteins
Subjects: UNSPECIFIED
Divisions: Div B > Solid State Electronics and Nanoscale Science
Depositing User: Cron Job
Date Deposited: 07 Mar 2014 11:41
Last Modified: 14 Dec 2014 17:46
DOI: 10.1098/rstb.2011.0410