The Saccharomyces cerevisiae URA7-encoded CTP synthetase is phosphorylated and stimulated by protein kinases A and C. Previous studies have revealed that Ser424 is the target site for protein kinase A. Using a purified S424A mutant CTP synthetase enzyme, we examined the effect of Ser424 phosphorylation on protein kinase C phosphorylation. The S424A mutation in CTP synthetase caused a 50% decrease in the phosphorylation of the enzyme by protein kinase C and an 80% decrease in the stimulatory effect on CTP synthetase activity by protein kinase C. The S424A mutation caused increases in the apparent Km values of CTP synthetase and ATP of 20-and 2-fold, respectively, in the protein kinase C reaction. The effect of the S424A mutation on the phosphorylation reaction was dependent on time and protein kinase C concentration. A CTP synthetase synthetic peptide (SLGRKDSHSA) containing Ser424 was a substrate for protein kinase C. Comparison of phosphopeptide maps of the wild type and S424A mutant CTP synthetase enzymes phosphorylated by protein kinases A and C indicated that Ser424 was also a target site for protein kinase C. Phosphorylation of Ser424 accounted for 10% of the total phosphorylation of CTP synthetase by protein kinase C. The incorporation of [methyl-3H]choline into phosphocholine, CDP-choline, and phosphatidylcholine in cells carrying the S424A mutant CTP synthetase enzyme was reduced by 48, 32, and 46%, respectively, when compared with control cells. These data indicated that phosphorylation of Ser424 by protein kinase A or by protein kinase C was required for maximum phosphorylation and stimulation of CTP synthetase and that the phosphorylation of this site played a role in the regulation of phosphatidylcholine synthesis by the CDP-choline pathway.
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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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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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