Prion-impairing mutations in Hsp70 chaperone Ssa1: effects on ATPase and chaperone activities.

Archives of Biochemistry and Biophysics
Patrick G Needham, Daniel C Masison

Abstract

We previously described many Hsp70 Ssa1p mutants that impair [PSI(+)] prion propagation in yeast without affecting cell growth. To determine how the mutations alter Hsp70 we analyzed biochemically the substrate-binding domain (SBD) mutant L483W and the nucleotide-binding domain (NBD) mutants A17V and R34K. Ssa1(L483W) ATPase activity was elevated 10-fold and was least stimulated by substrates or Hsp40 co-chaperones. Ssa1(A17V) and Ssa1(R34K) ATPase activities were nearly wild type but both showed increased stimulation by substrates. Peptide binding and reactivation of denatured luciferase were enhanced in Ssa1(A17V) and Ssa1(R34K) but compromised in Ssa1(L483W). The nucleotide exchange factor Fes1 influenced ATPase of wild type Ssa1 and each mutant differently. Partial protease digestion uncovered similar and distinct conformational changes of the substrate-binding domain among the three mutants. Our data suggest that prion-impairing mutations of Ssa1 can increase or decrease substrate interactions, alter the Hsp70 reaction cycle at different points and impair normal NBD-SBD cooperation.

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Citations

Sep 14, 2010·The Protein Journal·Yusuf TutarLütfi Tutar
Aug 3, 2011·Proceedings of the National Academy of Sciences of the United States of America·Deepak Sharma, Daniel C Masison
Mar 12, 2011·Journal of Molecular Biology·Gary P NewnamYury O Chernoff
Jul 26, 2014·Molecular Microbiology·Kevin C Stein, Heather L True
Apr 29, 2015·Journal of Molecular Biology·Patrick G NeedhamJeffrey L Brodsky
Jun 27, 2009·Prion·Daniel C MasisonDeepak Sharma
May 9, 2009·Journal of Molecular Biology·Vidhu MathurSusan W Liebman
Apr 18, 2009·Molecular Aspects of Medicine·Tobias JungTilman Grune
Nov 11, 2017·Cellular and Molecular Life Sciences : CMLS·Linan XuGary W Jones
Mar 25, 2020·Molecular and Cellular Biology·Chung-Hsuan KaoTanya T Paull

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