RGD Reference Report - Characterization of the phosphatase activity of a baculovirus-expressed calcineurin A isoform. - Rat Genome Database

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Characterization of the phosphatase activity of a baculovirus-expressed calcineurin A isoform.

Authors: Perrino, BA  Fong, YL  Brickey, DA  Saitoh, Y  Ushio, Y  Fukunaga, K  Miyamoto, E  Soderling, TR 
Citation: Perrino BA, etal., J Biol Chem 1992 Aug 5;267(22):15965-9.
RGD ID: 727532
Pubmed: PMID:1322410   (View Abstract at PubMed)

Calcineurin A was purified by calmodulin-Sepharose affinity chromatography from Sf9 cells infected with recombinant baculovirus containing the cDNA of a rat calcineurin A isoform. The Sf9-expressed calcineurin A has a low basal phosphatase activity in the presence of EDTA (0.9 nmol/min/mg) which is stimulated 3-5-fold by Mn2+. Calmodulin increased the Mn2+ stimulated activity 3-5-fold. Bovine brain calcineurin B increased the A subunit activity 10-15-fold, and calmodulin further stimulated the activity of reconstituted A and B subunits 10-15-fold (644 nmol/min/mg). The Km of calcineurin A for 32P-RII pep (a peptide substrate (DLDVPIPGRFDRRVSVAAE) for CaN), was 111 microM with or without calmodulin, and calmodulin increased the Vmax about 4-fold. The Km of reconstituted calcineurin A plus B for 32P-RII pep was 20 microM, and calmodulin increased the Vmax 18-fold without affecting the Km. CaN A467-492, a synthetic autoinhibitory peptide (ITSFEEAKGLDRINERMPPRRDAMP) from calcineurin, inhibited the Mn2+/calmodulin-stimulated activities of the reconstituted enzyme and the A subunit with IC50's of 25 microM and 90 microM, respectively. The reconstitution of the phosphatase activity of an expressed isoform of calcineurin A by purified B subunit and calmodulin may facilitate comparative studies of the regulation of calcineurin A activity by the B subunit and calmodulin.

Gene Ontology Annotations    Click to see Annotation Detail View

Biological Process
TermQualifierEvidenceWithReferenceNotesSourceOriginal Reference(s)
protein dephosphorylation  IDA 727532 RGD 

Molecular Function
TermQualifierEvidenceWithReferenceNotesSourceOriginal Reference(s)
phosphoprotein phosphatase activity  IDA 727532 RGD 

Objects Annotated

Genes (Rattus norvegicus)
Ppp3ca  (protein phosphatase 3 catalytic subunit alpha)

Additional Information