Substrate specificity of the nuclear protein kinase NII from porcine liver. Studies with casein variants.

The substrate specificity of the nuclear protein kinase NII from pig liver was studied using purified casein variants alpha s1B and beta A2 both in the phosphorylated and dephosphorylated form. Chymotryptic peptides of the phosphorylated casein were analyzed by ascending thin-layer chromatography followed by thin-layer electrophoresis. Phosphorylated peptides were detected by autoradiography. The determination of a phosphorylated site in a cluster of three serinees was performed by solid phase Edman degradation of the whole beta A2 casein. Phosphorylated beta A2 casein was by far the best substrate. A threonine at position 41 in the amino acid sequence was found to be predominantly phosphorylated. Dephosphorylated beta A2 casein became rephosphorylated at serine-17 whereas phosphorylated at threonine-41 was greatly reduced. In both sites acidic amino acid side chains are present at the C-terminal side though at different positions suggesting that acidic residues in the vicinity of the serin or threonine residue to be phosphorylated play an important role in the recognition by the nuclear protein kinase NII.