Protein geranylgeranyltransferase type-I (PGGTase-I) catalyzes alkylation of the cysteine residue in proteins containing a consensus C-terminal CaaX sequence ending in Leu or Phe by the C20 hydrocarbon moiety in geranylgeranyl diphosphate (GGPP). A kinetic study of the alkylation reaction was conducted with a continuous assay based on the fluorescence enhancement that accompanies geranylgeranylation of dansyl-GCIIL. The kinetic constants k(cat) = 0.34 +/- 0.01 s(-1), K(M)(G) = 0.86 +/- 0.05 microM for GGPP, and K(M)(D) = 1.6 +/- 0.1 microM for dansyl-GCIIL were calculated from initial rates measured at varying concentrations of the substrates. Inhibitor studies were conducted with dead-end inhibitors for GGPP and the peptide substrate. Double reciprocal plots for the peptide mimic Cys-AMBA-Leu gave a competitive pattern when plotted against varying concentrations of dansyl-GCIIL and an uncompetitive pattern against GGPP. Similar plots for 1-phosphono-(E,E,E)-geranylgeraniol, a dead-end inhibitor for GGPP, gave a competitive double reciprocal plot for varied concentrations of GGPP and induced potent substrate inhibition by dansyl-GCIIL when dansyl-GCIIL was the varied substrate. The dissociation constant (K(D)) for the PGGTase-I x GGPP complex was 120 +/- 20 nM. These results are consistent with an ordered binding mechanism for PGGTase-I where GGPP adds before peptide.
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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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