title: Identification of a latent Ca2+/calmodulin dependent protein kinase II phosphorylation site in vascular calpain II.
abstract: The Ca(2+)-dependent protease, calpain II, isolated from vascular smooth muscle was found to be a substrate for Ca2+/calmodulin-dependent protein kinase II (CaM kinase II) in vitro. Phosphorylation was dependent upon prior autolysis of the regulatory subunit of calpain II. The stoichiometry of phosphorylation of native, unautolyzed calpain II was 0.02 +/- 0.01 mol PO4/mol enzyme while for autolyzed calpain, the stoichiometry was 1.04 +/- 0.15 mol PO4/mol enzyme. All phosphate was incorporated into the 76 kDa catalytic subunit of calpain II. A single serine residue in domain III of the catalytic subunit was identified as the phosphate acceptor: RGS*TAGGCR. Phosphorylation doubled enzyme activity measured both as proteolysis of an exogenous substrate (alpha-casein) as well as by intermolecular catalytic subunit autolysis. The effects of phosphorylation could be reversed by dephosphorylation using a type IIA phosphoprotein phosphatase. These results demonstrate that calpain II possesses a latent CaM kinase II phosphorylation site that is unmasked by autolysis.
authors.affiliation: Krannert Institute of Cardiology, Indiana University School of Medicine, Indianapolis 46202-4800.
authors.forename: P
authors.initials: P
authors.lastname: McClelland
authors.forename: L P
authors.initials: LP
authors.lastname: Adam
authors.forename: D R
authors.initials: DR
authors.lastname: Hathaway
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000378:metabolism*; D002154:Calpain / Q000378:metabolism*; D002417:Cattle; D066298:In Vitro Techniques; D008969:Molecular Sequence Data; D009131:Muscle, Smooth, Vascular / Q000201:enzymology*; D010766:Phosphorylation
chemical_list: D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D002154:Calpain