Soluble cAMP-independent protein kinase from human spleen

Abstract

A protein kinase (EC 2.7.1.37) was purified 2000-fold, from the soluble protein fraction of human spleen cells, using ion-exchange chromatography, ammonium sulfate fractionation, and gel filtration. This rapid procedure yielded 30% of the initial activity and an enzyme preparation with specific activity of 62 nmol min −1 mg −1 of protein. On the basis of disc gel electrophoresis in sodium dodecyl sulfate acrylamide gels and isoelectric focusing the enzyme preparation appears homogeneous and to consist of one polypeptide with a molecular weight of 43,000 and having a p I of 7.1. The purified enzyme activity is cyclic AMP and cGMP independent phosphorylates both α-casein and phosvitin, and uses Mg 2+ ATP and Mg 2+ GTP as phosphate donors, exhibiting an apparent K m of 2.0 and 6.6 × 10 −5 m, respectively. Furthermore, the enyzme activity is strongly inhibited by heparin ( K 50 = 0.1 μg/ml). These catalytic properties are characteristic of the enzyme casein kinase II, as described in several eukaryotic cells.