Abstract
To determine the activity of all human Cdc25 phosphatases, two different methods are described. For assaying phosphatase activities of recombinant Cdc25 proteins produced in Escherichia coli or insect cells, a fluorimetric assay using fluorescein diphosphate (FDP) as a substrate is recommended. To analyze endogenous Cdc25 phosphatase activities of immunoprecipitates from total cellular extracts, the physiological substrate Cdk1/cyclin B1 is most sensitive.
Publication types
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Research Support, U.S. Gov't, P.H.S.
MeSH terms
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Animals
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CDC2 Protein Kinase / metabolism*
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Cyclin B / metabolism
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Cyclin B1
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Escherichia coli / metabolism
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Fluorescein / metabolism
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Glutathione Transferase / genetics
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Glutathione Transferase / metabolism
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HeLa Cells
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Humans
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Phosphorylation
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Protein Kinases / metabolism*
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Recombinant Fusion Proteins / analysis
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S Phase
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Spodoptera / cytology
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Spodoptera / genetics
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cdc25 Phosphatases / analysis*
Substances
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CCNB1 protein, human
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Cyclin B
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Cyclin B1
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Recombinant Fusion Proteins
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Glutathione Transferase
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Protein Kinases
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histone H1 kinase
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CDC2 Protein Kinase
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cdc25 Phosphatases
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Fluorescein