title: Ca2+/calmodulin dependent protein kinase from Mycobacterium smegmatis ATCC 607.
abstract: A soluble Ca2+/calmodulin dependent protein kinase has been partially purified (approximately 400 fold) from Mycobacterium smegmatis ATCC 607 using several purification steps like ammonium sulphate precipitation (30-60%), Sepharose CL-6B gel filtration, DEAE-cellulose and finally calmodulin-agarose affinity chromatography. On SDS-PAGE, this enzyme preparation showed a major protein band of molecular mass 35 kD and its activity was dependent on calcium, calmodulin and ATP when measured under saturating histone IIs (exogenous substrate) concentration. Phosphorylation of histone IIs was inhibited by W-7 (calmodulin inhibitor) and KN-62 (CaM-kinase inhibitor) with IC50 of 1.5 and 0.25 microm respectively, but was not affected by inhibitors of PKA (Sigma P5015) and PKC (H-7). All these results confirm that purified enzyme is Ca2+/calmodulin dependent protein kinase of M. smegmatis. The protein kinase of M. smegmatis demonstrated a narrow substrate specificity for both exogenous as well as endogenous substrates. These results suggest that purified CaM-kinase must be involved in regulating specific function(s) in this organism.
authors.affiliation: Department of Biochemistry, Postgraduate Institute of Medical Education and Research, Chandigarh, India.
authors.forename: S
authors.initials: S
authors.lastname: Sharma
authors.forename: S
authors.initials: S
authors.lastname: Giri
authors.forename: G K
authors.initials: GK
authors.lastname: Khuller
mesh_terms: D000255:Adenosine Triphosphate / Q000502:physiology; D001345:Autoradiography; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000302:isolation & purification* / Q000378:metabolism; D002147:Calmodulin / Q000037:antagonists & inhibitors / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D004795:Enzyme Stability; D007700:Kinetics; D009161:Mycobacterium / Q000201:enzymology*; D010766:Phosphorylation; D013379:Substrate Specificity
chemical_list: D002147:Calmodulin; D000255:Adenosine Triphosphate; D017871:Calcium-Calmodulin-Dependent Protein Kinases