The Hsp104 protein from Saccharomyces cerevisiae is a member of the Hsp100/Clp family of molecular chaperones. It mediates the solubilization of aggregated proteins in an ATP-dependent process assisted by the Hsp70/40 system. Although the principal function of Hsp104 is well established, the mechanistic details of this catalyzed disaggregation are poorly understood. In this work, we have investigated the interaction of Hsp104 with reduced, carboxymethylated alpha-lactalbumin (RCMLa), a permanently unfolded model substrate. Our results demonstrate that the affinity of Hsp104 toward polypeptides is regulated by nucleotides. In the presence of ATP or adenosine-5' -O-(3-thiotriphosphate), the chaperone formed complexes with RCMLa, whereas no binding was observed in the presence of ADP. In particular, the occupation of the N-terminally located nucleotide-binding domain with ATP seems to be crucial for substrate interaction. When ATP binding to this domain was impaired by mutation, Hsp104 lost its ability to interact with RCMLa. Our results also indicate that upon association with a polypeptide, a conformational change occurs within Hsp104 that strongly reduces the dynamics of nucleotide exchange and commits the bound polypeptide to ATP hydrolysis.
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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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Evidence ID | Analyze ID | File | Description |
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