title: Biochemical characterization of a recombinant Swainsona canescens calcium-dependent protein kinase (ScCPK1).
abstract: Calcium-dependent protein kinases (CPKs) constitute a unique family of kinases involved in many physiological responses in plants. Biochemical and kinetic properties of a recombinant Swainsona canescens calcium-dependent protein kinase (ScCPK1) were examined in this study. The optimum pH and temperature for activity were pH 7.5 and 37 °C, respectively. Substrate phosphorylation activity of ScCPK1 was calmodulin (CaM) independent. Yet CaM antagonists, W7 [N-(6-aminohexyl)-5-chloro-1-naphthalene sulphonamide] and calmidazolium inhibited the activity with IC(50) values of 750 nM and 350 μM, respectively. Both serine and threonine residues were found to be phosphorylated in autophosphorylated ScCPK1 and in histone III-S phosphorylated by ScCPK1. The [Ca(2)(+)] for half maximal activity (K(0.5)) was found to be 0.4 μM for ScCPK1 with histone III-S as substrate. Kinetic analysis showed that K(M) of ScCPK1 for histone III-S was 4.8 μM. These data suggest that ScCPK1 is a functional Ser/Thr kinase, regulated by calcium, and may have a role in Ca(2)(+)-mediated signaling in S. canescens.
authors.affiliation: Department of Biochemistry, Indian Institute of Science, Bangalore 560012, India.
authors.forename: S M
authors.initials: SM
authors.lastname: Srideshikan
authors.forename: V L
authors.initials: VL
authors.lastname: Vanishree
authors.forename: C
authors.initials: C
authors.lastname: Jayabaskaran
mesh_terms: D002118:Calcium / Q000378:metabolism*; D002147:Calmodulin / Q000037:antagonists & inhibitors / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D007887:Fabaceae / Q000201:enzymology* / Q000378:metabolism; D006657:Histones / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007093:Imidazoles / Q000494:pharmacology; D010766:Phosphorylation; D010940:Plant Proteins / Q000378:metabolism*; D011494:Protein Kinases / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D012694:Serine / Q000378:metabolism*; D015398:Signal Transduction; D013449:Sulfonamides / Q000494:pharmacology; D013696:Temperature; D013912:Threonine / Q000378:metabolism*; D017871:Calcium-Calmodulin-Dependent Protein Kinases
chemical_list: D002118:Calcium; D002147:Calmodulin; D004791:Enzyme Inhibitors; D006657:Histones; D007093:Imidazoles; D010940:Plant Proteins; D011494:Protein Kinases; D011994:Recombinant Proteins; D012694:Serine; D013449:Sulfonamides; D013912:Threonine; D017871:Calcium-Calmodulin-Dependent Protein Kinases; C031938:calmidazolium; C017967:W 7