A novel endogenous PP2C-like phosphatase dephosphorylates casein kinase II-phosphorylated Physarum fragmin.
Plasmodial fragmin, a Physarum polycephalum F-actin severing and capping protein, is phosphorylated by casein kinase II at Ser(266) (De Corte, V., Gettemans, J., De Ville, Y., Waelkens, E., and Vandekerckchove, J. (1996), Biochemistry 35, 5472-5480). In this study, we report the purification and characterization of the corresponding fragmin phosphatases. One of the enzymes was purified to near homogeneity from a cytosolic extract; it dephosphorylates CKII-phosphorylated fragmin, a peptide encompassing the CKII phosphorylation site of fragmin as well as histone 2A, CKII-phosphorylated casein and the CKII model-peptide substrate: R(3)E(3)S(P)E(3). Its activity was highly stimulated by Mn(2+) and Mg(2+), and based on its lack of sensitivity toward phosphatase effectors we could exclude similarities with PP1, PP2A and PP2B phosphatases. All biochemical properties of the phosphatase point to a PP2C-like enzyme. A second phosphatase dephosphorylating fragmin was identified as a Physarum alkaline phosphatase.
Waelkens
E
E
Division of Biochemistry, Katholieke Universiteit Leuven, Leuven, B-3000, Belgium.
de Corte
V
V
Merlevede
W
W
Vandekerckhove
J
J
Gettemans
J
J
D000469:Alkaline Phosphatase / Q000302:isolation & purification / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D016718:Arachidonic Acid / Q000494:pharmacology; D002352:Carrier Proteins / Q000378:metabolism*; D047390:Casein Kinase II; D002848:Chromatography, DEAE-Cellulose; D007700:Kinetics; D008274:Magnesium / Q000494:pharmacology; D008345:Manganese / Q000494:pharmacology; D008840:Microfilament Proteins / Q000378:metabolism*; D008969:Molecular Sequence Data; D010749:Phosphoprotein Phosphatases / Q000302:isolation & purification / Q000378:metabolism*; D016839:Physarum polycephalum / Q000201:enzymology*; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D012694:Serine; D013379:Substrate Specificity
D002352:Carrier Proteins; C034037:F-actin-binding proteins; D008840:Microfilament Proteins; D016718:Arachidonic Acid; D008345:Manganese; D012694:Serine; D047390:Casein Kinase II; D017346:Protein Serine-Threonine Kinases; D000469:Alkaline Phosphatase; D010749:Phosphoprotein Phosphatases; D008274:Magnesium