The yeast ubiquitin (Ub) conjugating enzyme CDC34 plays a crucial role in the progression of the cell cycle from the G1 to S phase. In an effort to identify proteins that interact with CDC34 we undertook a genetic screen to isolate genes whose increased expression suppressed the cell cycle defect associated with the cdc34-2 temperature-sensitive allele. From this screen, the poly-Ub gene UBI4 was identified as a moderately strong suppressor. The fact that the overexpression of a gene encoding a single Ub protein could also suppress the cdc34-2 allele indicated that suppression was related to the increased abundance of Ub. Ub overexpression was found to suppress two other structurally unrelated cdc34 mutations, in addition to the cdc34-2 allele. In all three cases, suppression depended on the expression of Ub with an intact carboxyl terminus. Only the cdc34-2 allele, however, could be suppressed by Ub with an amino acid substitution at lysine 48 which is known to be involved in multi-Ub chain assembly. Genetic results showing allele specific suppression of cdc34 mutations by various Ub derivatives suggested a specific noncovalent interaction between Ub and CDC34. Consistent with this prediction, we have shown by chemical cross-linking the existence of a specific noncovalent Ub binding site on CDC34. Together, these genetic and biochemical experiments indicate that Ub suppression of these cdc34 mutations results from the combined contributions of Ub-CDC34 thiol ester formation and a noncovalent interaction between Ub and CDC34 and therefore suggest that the correct positioning of Ub on a surface of the ubiquitin conjugating enzyme is a requirement of enzyme function.
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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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