Purification and properties of cAMP-independent nuclear protein kinase from Dictyostelium discoideum.
A cyclic-AMP-independent nuclear protein kinase has been purified from Dictyostelium discoideum amoebae. The purification procedure involves chromatography of DEAE-Sephadex, phosphocellulose and heparin-Sepharose. The purified enzyme phosphorylates threonine and serine of acidic proteins as casein and phosvitin. Phosphorylation of casein is stimulated by spermine. The kinase requires Mg2+ and can utilize both ATP and GTP as phosphoryl donors. Heparin is a potent inhibitor of the enzyme, being the protein kinase activity fully inhibited at concentrations of 0.5 micrograms/ml. One polypeptide of molecular mass 38 kDa was the major protein band present in the purified kinase preparation as estimated by NaDodSO4 denaturing polyacrylamide gel electrophoresis. This band belongs to the protein kinase because it is the only one that is observed associated with the protein kinase activity when the enzyme preparation is centrifuged in glycerol gradients. The 38-kDa polypeptide is also the major product of autophosphorylation of the enzyme preparation. The enzymatic properties allow to classify the enzyme as a type-II casein kinase. However, its structural properties are different from the mammalian type-II casein kinases and make the D. discoideum enzyme more similar to the plants type-II casein kinases.
D000255:Adenosine Triphosphate / Q000378:metabolism; D002467:Cell Nucleus / Q000201:enzymology*; D002499:Centrifugation, Density Gradient; D002845:Chromatography / Q000379:methods; D000242:Cyclic AMP / Q000494:pharmacology; D004023:Dictyostelium / Q000201:enzymology*; D004591:Electrophoresis, Polyacrylamide Gel; D005786:Gene Expression Regulation; D006160:Guanosine Triphosphate / Q000378:metabolism; D006493:Heparin / Q000494:pharmacology; D008670:Metals / Q000502:physiology; D008970:Molecular Weight; D011073:Polyamines / Q000494:pharmacology; D011494:Protein Kinases / Q000302:isolation & purification* / Q000378:metabolism; D013379:Substrate Specificity
D008670:Metals; D011073:Polyamines; D006160:Guanosine Triphosphate; D000255:Adenosine Triphosphate; D006493:Heparin; D000242:Cyclic AMP; D011494:Protein Kinases