Cak1p is an essential protein kinase that phosphorylates and thereby activates the major cyclin-dependent kinase in budding yeast, Cdc28p. The sequence of Cak1p differs from other members of the protein kinase superfamily in several conserved regions. Cak1p lacks the highly conserved glycine loop motif (GXGXXG) that is found in the nucleotide binding fold of virtually all protein kinases and also lacks a number of conserved amino acids found at sites throughout the protein kinase core sequence. We have used kinetic and mutagenic analyses to investigate whether these sequence differences affect the nucleotide-binding properties of Cak1p. Although Cak1p differs dramatically from other protein kinases, it binds ATP with a reasonable affinity, with a KM of 4.8 microM. Mutations of the putative invariant lysine in Cak1p (Lys-31), homologous to a residue required for activity in virtually all protein kinases and that interacts with the ATP phosphates, moderately reduced the ability of Cak1p to bind ATP but did not dramatically affect the catalytic rate of the kinase. Similarly, Cak1p is insensitive to the ATP analog 5'-fluorosulfonylbenzoyladenosine, which inhibits most protein kinases through covalent modification of the invariant lysine. We found that Cak1p is tolerant of mutations within its glycine loop region. Remarkably, Cak1p remains functional even following truncation of its first 31 amino acids, including the glycine loop region and the invariant lysine. We conclude that the Cak1p nucleotide-binding pocket differs significantly from those of most other protein kinases and therefore might provide a specific target for an inhibitory drug.
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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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