Studies on the phosphorylation of a M(r) 25,000 protein, a putative protein phosphatase 2A modulator, by casein kinase I, and analysis of multiple endogenous phosphates.
A M(r) 25,000 protein, which was isolated from the cytosolic fraction of Xenopus laevis oocytes, is a newly identified substrate for casein kinase II and protein kinase C [Hashimoto et al. (1995) J. Biochem. 118, 453-460], and was recently shown to have the ability to modulate protein phosphatase 2A activity [Hashimoto et al. (1996) J. Biochem. 119, 626-632]. Acid phosphatase treatment of the protein shifted its electrophoretic mobility from 25 to 20 kDa on SDS-PAGE. The content of alkali-labile phosphate bound covalently to the protein was 53 mol per mol of M(r) 25,000 protein. Amino acid composition analysis revealed that there are 50 serine residues and 6 threonine residues per mol of this protein. Therefore, this M(r) 25,000 protein seems to be highly phosphorylated in vivo. The M(r) 25,000 protein, once partially dephosphorylated by acid phosphatase, served as an efficient substrate for casein kinase I and casein kinase II. When entirely dephosphorylated, the M(r) 25,000 protein was used as a substrate, the rate of phosphorylation with both casein kinases being decreased. This behavior of casein kinases toward the M(r) 25,000 protein reflects the possible mechanism of multisite phosphorylation in which the introduction of a phosphate group facilitates sequential phosphorylation.
Liu
Y
Y
Department of Pathological Biochemistry, School of Life Sciences, Faculty of Medicine, Tottori University.
Kohara
H
H
Hashimoto
E
E
D000135:Acid Phosphatase / Q000378:metabolism; D000818:Animals; D047390:Casein Kinase II; D047388:Casein Kinases; D004591:Electrophoresis, Polyacrylamide Gel; D006493:Heparin / Q000494:pharmacology; D007546:Isoquinolines / Q000494:pharmacology; D007700:Kinetics; D008970:Molecular Weight; D019319:Okadaic Acid / Q000494:pharmacology; D009865:Oocytes; D010749:Phosphoprotein Phosphatases / Q000378:metabolism; D010750:Phosphoproteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D010761:Phosphorus Radioisotopes; D010766:Phosphorylation; D047428:Protein Kinase Inhibitors; D011494:Protein Kinases / Q000378:metabolism*; D054648:Protein Phosphatase 2; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D011506:Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D014982:Xenopus laevis; D000097847:Protein Phosphatase Inhibitory Proteins
D000135:Acid Phosphatase; D047390:Casein Kinase II; D047388:Casein Kinases; D006493:Heparin; D007546:Isoquinolines; D019319:Okadaic Acid; D010749:Phosphoprotein Phosphatases; D010750:Phosphoproteins; D010761:Phosphorus Radioisotopes; D047428:Protein Kinase Inhibitors; D011494:Protein Kinases; D054648:Protein Phosphatase 2; D017346:Protein Serine-Threonine Kinases; D011506:Proteins; C024833:protein phosphatase inhibitor-2; D000097847:Protein Phosphatase Inhibitory Proteins; C058815:N-(2-aminoethyl)-5-chloroisoquinoline-8-sulfonamide