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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| Evidence ID | Analyze ID | Gene/Complex | Systematic Name/Complex Accession | Qualifier | Gene Ontology Term ID | Gene Ontology Term | Aspect | Annotation Extension | Evidence | Method | Source | Assigned On | Reference |
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| Evidence ID | Analyze ID | Gene | Gene Systematic Name | Phenotype | Experiment Type | Experiment Type Category | Mutant Information | Strain Background | Chemical | Details | Reference |
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| Evidence ID | Analyze ID | Gene | Gene Systematic Name | Disease Ontology Term | Disease Ontology Term ID | Qualifier | Evidence | Method | Source | Assigned On | Reference |
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| Evidence ID | Analyze ID | Regulator | Regulator Systematic Name | Target | Target Systematic Name | Direction | Regulation of | Happens During | Regulator Type | Direction | Regulation Of | Happens During | Method | Evidence | Strain Background | Reference |
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| Site | Modification | Modifier | Source | Reference |
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| Evidence ID | Analyze ID | Interactor | Interactor Systematic Name | Interactor | Interactor Systematic Name | Allele | Assay | Annotation | Action | Phenotype | SGA score | P-value | Source | Reference | Note |
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| Evidence ID | Analyze ID | Interactor | Interactor Systematic Name | Interactor | Interactor Systematic Name | Assay | Annotation | Action | Modification | Source | Reference | Note |
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| Complement ID | Locus ID | Gene | Species | Gene ID | Strain background | Direction | Details | Source | Reference |
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| Evidence ID | Analyze ID | Dataset | Description | Keywords | Number of Conditions | Reference |
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