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At least 19 recordsLinked to original sources
PubMed1991

Purification and characterization of polyamine-stimulated protein kinase (casein kinase II) from bovine spermatozoa.

Casein kinase II from bovine epididymal spermatozoa was purified to apparent homogeneity by repeated chromatography with phosphocellulose and gel filtration with sephacryl S-200. The purified enzyme exhibited a molecular mass of 130 kDa by gel filtration and displayed three polypeptide bands with molecular masses of 26, 33, and 36 kDa by SDS-polyacrylamide gel electrophoresis. Antibodies raised against calf thymus casein kinase II cross reacted with the three sperm polypeptides. Incubation of the holoenzyme with either [gamma-32P]ATP or [gamma-32P]GTP resulted in the phosphorylation of the 26-kDa subunit. The enzymatic activity with casein as substrate was strongly inhibited by nanomolar heparin and greatly stimulated by micromolar spermine. With casein as substrate, the specific activity of the pure enzyme (0.5 mumol/min/mg protein) was comparable to that of casein kinase II from other sources. Endogenous substrates of the kinase were demonstrated by incubating sperm cytosolic extracts with [gamma-32P]GTP, under conditions that limit the expression of other protein kinases, and analyzing the products by SDS-PAGE and autoradiography. Similar results were obtained when sperm extracts, suitably diluted to minimize endogenous casein kinase II, were incubated with [gamma-32P]GTP and aliquots of pure sperm casein kinase II. Low concentrations (50 microM) spermine strongly enhanced the phosphorylation of 92- and 106-kDa cytosolic proteins. Our results clearly show that casein kinase II is present in spermatozoa and that it shares many of the properties of the enzyme from other sources. Further, they indicate that the enzyme plays a role in mediating the phosphorylation state of sperm proteins.

Animals↗
PubMed1995

Direct identification of a polyamine binding domain on the regulatory subunit of the protein kinase casein kinase 2 by photoaffinity labeling.

Phosphorylation of many protein substrates by the protein kinase casein kinase 2 (CK2) is stimulated severalfold in the presence of polyamines such as spermine. Previous experiments have shown that CK2 is a polyamine binding protein and that the regulatory beta subunit is required for this binding activity. To delineate the spermine binding site of CK2, we have applied a photoaffinity labeling method using a tritiated photoactivable analog of spermine, [3H]sperminediazonium. The photoaffinity labeled beta subunit was cleaved with cyanogen bromide, and two labeled peptides were separated by high performance liquid chromatography. The major one was the peptide T72EQAAEM78 and the minor one was a 22-amino acid peptide comprising residues Ile98 to Met119. Thr72 and His108 were identified as the labeled amino acids of the Thr72-Met78 and Ile98-Met119 peptides, respectively. In the same manner, we succeeded in determining the residue Leu220 as an alpha subunit residue covalently bound to the probe. The photoaffinity labeling method described here enabled the first elucidation, by direct microsequencing, of a polyamine binding site on CK2 for which we propose a provisional structural model. These observations suggest a possible mechanism for CK2 activation by polyamines at the molecular level.

Affinity Labels↗
PubMed1995

Differential phosphorylation of ribosomal acidic proteins from yeast cell by two endogenous protein kinases: casein kinase-2 and 60S kinase.

The native 80S ribosomes isolated from Saccharomyces cerevisiae (strain W303) cells was phosphorylated by two endogenous protein kinases: multifunctional casein kinase-2 (CK-2) and specific 60S kinase. Three acidic proteins within the 60S ribosomal subunit: YP1 beta, YP1 beta' and YP2 alpha are phosphorylated by both kinases. The other two proteins: YP1 alpha and YP2 beta are predominantly phosphorylated by CK-2 but not by 60S kinase. This was confirmed in the experiment with the recombinant protein, YP2 beta, as a substrate, which is practically not phosphorylated by specific 60S kinase. These results together with the previous data based on the target amino-acid sequences suggest that, in addition to the multifunctional casein kinase-2 and specific 60S kinase, there exist probably other protein kinase(s) which phosphorylate the ribosomal acidic proteins in the cell.

Casein Kinase II↗
PubMed1993

Control of simian virus 40 DNA replication by the HeLa cell nuclear kinase casein kinase I.

The initiation of simian virus 40 (SV40) DNA replication is regulated by the phosphorylation state of the viral initiator protein, large T antigen. We describe the purification from HeLa cell nuclei of a 35-kDa serine/threonine protein kinase that phosphorylates T antigen at sites that are phosphorylated in vivo and thereby inhibits its ability to initiate SV40 DNA replication. The inhibition of both origin unwinding and DNA replication by the kinase is reversed by protein phosphatase 2A. As determined by molecular weight, substrate specificity, autophosphorylation, immunoreactivity, and limited sequence analysis, this kinase appears to be identical to casein kinase I, a ubiquitous serine/threonine protein kinase that is closely related to a yeast kinase involved in DNA metabolism. The HeLa cell phosphorylation cycle that controls the initiation of SV40 DNA replication may also play a role in cellular DNA metabolism.

Adenosine Triphosphate↗
PubMed1984

Some properties of the nucleotide-binding site of troponin T kinase-casein kinase type II from skeletal muscle.

Investigation of properties of skeletal muscle troponin T kinase (EC 2.7.1.37) has revealed that the enzyme belongs to the group of casein kinases of the second type. The enzyme consists of two subunits with apparent molecular weights of 44 000 and 26 000 and contains a protein with molecular weight of 39 000, which is probably the proteolytic fragment of the 44 000 subunit. The substrate specificity of troponin T kinase was tested, using 20 analogs of the nucleotide. The enzyme has a low substrate specificity toward the purine base and uses both ATP and GTP as substrates. Modification of the ribose ring does not influence the enzyme interaction with the nucleotide; however, the cleavage of ribose leads to a decrease of the enzyme-nucleotide interaction. Elimination of the gamma-terminal phosphate or its modification by bulky hydrophobic radicals do not affect this interaction. A comparison of the Ki values for different analogs suggests that the interaction of troponin T kinase with the nucleotide occurs via the binding of the purine base and the beta-phosphate group of the analog.

Adenosine Triphosphate↗
PubMed1985

Protein phosphatases active on acetyl-CoA carboxylase phosphorylated by casein kinase I, casein kinase II and the cAMP-dependent protein kinase.

The protein phosphatases in rat liver cytosol, active on rat liver acetyl-CoA carboxylase (ACC) phosphorylated by casein kinase I, casein kinase II and the cAMP-dependent protein kinase, have been partially purified by anion-exchange and gel filtration chromatography. The major phosphatase activities against all three substrates copurify through fractionation and appear to be identical to protein phosphatases 2A1 and 2A2. No unique protein phosphatase active on 32P-ACC phosphorylated by the casein kinases was identified.

Acetyl-CoA Carboxylase↗
PubMed1992

Determinants on simian virus 40 large T antigen are important for recognition and phosphorylation by casein kinase I.

Casein kinase I has been shown to phosphorylate Ser123 and possibly Thr124, in simian virus 40 (SV40) large T antigen; the same sites are also modified in cultured cells incubated with 32Pi [Friedrich A. Grässer, Karl H. Scheidtmann, Polygena T. Tuazon, Jolinda A. Traugh & Gernot Walter (1988) Virology 165, 13-22]. The peptide, A-D-S-Q-H-S-T-P-P, which corresponds to the amino acid sequence 118-125 of SV40 large T antigen, was synthesized together with peptides containing changes in specific amino acid residues on either side of Ser123. These peptides were used as model substrates to determine the amino acids in the SV40 large T antigen important for recognition by casein kinase I. The native peptide identified above, with aspartate at the -4 position, was a poor substrate for casein kinase I in vitro. Peptides with acidic residues added at the -2 and -3 positions, preceding Ser123, were phosphorylated by casein kinase I with apparent Km values around 2 mM and Vmax values up to 500 pmol.min-1.ml-1. When acidic residues were added at both sides of the phosphorylatable serine, the peptide had a first-order rate constant over 20-fold higher than peptides with acidic amino acid residues at the N-terminus only; the apparent Km value was 0.65 mM with a Vmax of 2900 pmol.min-1.ml-1. The effects of modifying Ser120 to phosphoserine were examined by addition of a recognition sequence for the cAMP-dependent protein kinase prior to Ser120. Prior phosphorylation of the peptide at Ser120 lowered the apparent Km to 0.061 mM and increased the Vmax to 360 pmol.min-1.ml-1, a 50-fold decrease in Km for casein kinase I and a 6-fold increase in Vmax as compared to the non-phosphorylated peptide. This indicates that Ser120, which has been shown to be phosphorylated in vivo, provides an appropriate recognition determinant for casein kinase I.

Amino Acid Sequence↗
PubMed1989

Phosphorylation of varicella-zoster virus glycoprotein gpI by mammalian casein kinase II and casein kinase I.

Varicella-zoster virus (VZV) glycoprotein gpI is the predominant viral glycoprotein within the plasma membranes of infected cells. This viral glycoprotein is phosphorylated on its polypeptide backbone during biosynthesis. In this report, we investigated the protein kinases which participate in the phosphorylation events. Under in vivo conditions, VZV gpI was phosphorylated on its serine and threonine residues by protein kinases present within lysates of either VZV-infected or uninfected cells. Because this activity was diminished by heparin, a known inhibitor of casein kinase II, isolated gpI was incubated with purified casein kinase II and shown to be phosphorylated in an in vitro assay containing [gamma-32P]ATP. The same glycoprotein was phosphorylated when [32P]GTP was substituted for [32P]ATP in the protein kinase assay. We also tested whether VZV gpI was phosphorylated by two other ubiquitous mammalian protein kinases--casein kinase I and cyclic AMP-dependent kinase--and found that only casein kinase I modified gpI. When the predicted 623-amino-acid sequence of gpI was examined, two phosphorylation sites known to be optimal for casein kinase II were observed. Immediately upstream from each of the casein kinase II sites was a potential casein kinase I phosphorylation site. In summary, this study showed that VZV gpI was phosphorylated by each of two mammalian protein kinases (casein kinase I and casein kinase II) and that potential serine-threonine phosphorylation sites for each of these two kinases were present in the viral glycoprotein.

Amino Acids↗
PubMed1983

Thyroid hormone increases type I adenosine 3', 5'-monophosphate-dependent protein kinase and casein kinase activities in rat liver cytosol: analysis of protein kinases by polyacrylamide disc gel electrophoresis.

We investigated the action of thyroid hormone on each protein kinase in rat liver cytosol. Kinases were analyzed by polyacrylamide disc gel electrophoresis and isoelectric focusing in polyacrylamide gel. Polyacrylamide disc gel electrophoresis separated cAMP-dependent protein kinase type I (Rf = 0.35), type II (Rf = 0.44), their catalytic subunit (Rf = 0.26), and cAMP-independent protein kinase (Rf = 0.50). Casein kinase was detected at Rf = 0.37. In addition to the catalytic subunit with Rf = 0.26, another catalytic subunit was found at Rf = 0.44 when the cytosol was preincubated with cAMP. The administration of T3 (20 micrograms/100 g BW for 3 days) to hypothyroid rats increased enzyme activities of type I holoenzyme and casein kinase by 48%. Free catalytic subunit, separated from holoenzyme, had the same level of enzyme activity in both groups, suggesting greater endogenous dissociation of type I holoenzyme in hypothyroid rats. When heat-inactivated rat liver cytosol was used as substrate in the assay of protein kinase activity, the peak enzyme active in phosphorylating the cytosol corresponded to the casein kinase peak. Our data indicate that casein kinase is the main enzyme that mediates phosphorylation of endogenous proteins in rat liver cytosol, and that T3 treatment increases the activity of casein kinase and of type I cAMP-dependent protein kinase.

Animals↗
PubMed1983

Polyglutamyl peptides: a new class of inhibitors of type-2 casein kinases.

Casein kinase-TS (Ck-TS), a type-2 casein kinase purified from rat liver cytosol which phosphorylates seryl and threonyl residues N-terminal to acidic clusters, is specifically inhibited by polyglutamyl peptides which are ineffective both on type-1 casein kinase and on cAMP-dependent protein kinase. The inhibition is competitive toward the protein substrate and non-competitive toward ATP. Among the polyglutamates tested (Glu)70 is the most effective (Ki 0.11 microM). (Glu)10 and (Glu)5 are also inhibitors, though less powerful than (Glu)70, while (Glu)3, (Glu)2 and free glutamic acid up to 5 mM are ineffective. These results disclose the possibility that naturally occurring polypeptides containing long stretches of acidic residues may act as physiological inhibitors of type-2 casein kinases.

Adenosine Triphosphate↗
PubMed1994

Dual role of calsequestrin as substrate and inhibitor of casein kinase-1 and casein kinase-2.

Calsequestrin from different muscle tissues and species has been phosphorylated by casein kinase-1 and casein kinase-2, in the conditions previously reported by Cala and Jones (J. Biol. Chem. 266, 391-398, 1991). Results indicates that rabbit cardiac and skeletal calsequestrin and frog skeletal calsequestrin are phosphorylated by both casein kinase-1 and casein kinase-2, at variance with chicken skeletal calsequestrin which is a poor substrate for both enzymes. We also observed that chicken calsequestrin is able to inhibit phosphorylation of cardiac calsequestrin, as well as other specific substrates, when added together to the assay medium.

Amino Acid Sequence↗
PubMed1993

Amyloid beta-protein stimulates casein kinase I and casein kinase II activities.

Amyloid beta-protein (A beta) is the major protein of cerebrovascular and plaque amyloid in Alzheimer's disease (AD). Extensive evidence has demonstrated abnormal protein phosphorylation in this disease. We investigated the effect of synthetic A beta with the amino-acid sequence corresponding to cerebrovascular A beta and plaque A beta on the activities of casein kinase I (CK I) and casein kinase II (CK II). These enzymes were purified from bovine brain and casein was used as a substrate. A beta was found to stimulate markedly CK I- and CK II-mediated phosphorylation of casein in a concentration-dependent manner. The effect of plaque A beta was considerably higher than that of cerebrovascular A beta. Heparin, which is known to be a specific inhibitor of CK II, completely inhibited A beta-stimulated CK II activity. A beta itself was not a substrate for casein kinases. These findings were confirmed using other substrates for CK I and CK II. The experiments with synthetic CK II-substrate peptide (Leu-Glu-Leu-Ser-Asp-Asp-Asp-Asp-Glu) and the phosphorylation of erythrocyte membrane proteins by intrinsic membrane-bound CK I in erythrocytes showed marked stimulation in activities of casein kinases in the presence of A beta 1-40 or blocked A beta. We propose that A beta, by stimulating casein kinases, may contribute to abnormal protein phosphorylation in AD, in particular to increased phosphorylation of microtubule-associated proteins, leading to the neurofibrillary tangles formation and neurodegeneration in this disease. Interaction of A beta with protein kinases, thus, may characterize the beginning of the disease.

Amino Acid Sequence↗
PubMed1983

Identification of calcium-independent myosin kinase with casein kinase II.

A crude myosin fraction from bovine brain has been found to contain a Ca2+-independent myosin kinase that catalyzes the phosphorylation of 20,000-Da light chain of gizzard myosin. The myosin kinase has been separated from the myosin by Sepharose CL-4B gel filtration and purified further by chromatography on phosphocellulose, Sephacryl S-300, and hydroxylapatite. The myosin kinase was found to copurify with casein kinase II and show the same substrate specificity with the casein kinase. These results indicate that the myosin kinase is identical to casein kinase II. The purified myosin kinase catalyzed the preferential phosphorylation of the threonyl residues of 20,000-Da light chains of gizzard and brain myosins. The 17,000-Da light chains of these myosins and the mixed light chains of skeletal and cardiac muscle myosins were not phosphorylated by the enzyme to an appreciable extent.

Amino Acids↗
PubMed1989

Synergistic phosphorylation of rabbit muscle glycogen synthase by cyclic AMP-dependent protein kinase and casein kinase I. Implications for hormonal regulation of glycogen synthase.

The phosphorylation of rabbit skeletal muscle glycogen synthase by casein kinase I is markedly enhanced if the enzyme has previously been phosphorylated by cAMP-dependent protein kinase. The presence of phosphate in the primary cAMP-dependent protein kinase sites, sites 1a, 1b, and 2 (serine 7), increases the activity of casein kinase I toward residues in the vicinity of these sites. This synergistic phosphorylation correlates with potent inactivation of the glycogen synthase. Analysis of the NH2 terminus of the enzyme subunit indicated that phosphorylation at serine 7 caused serine 10 to become a preferred casein kinase I site and that phosphoserine can be an important recognition determinant for casein kinase I. This finding can also explain how epinephrine stimulation of skeletal muscle provokes significant increases in the phosphorylation state of serine residues, in particular serine 10, not recognized by cAMP-dependent protein kinase.

Amino Acid Sequence↗
PubMed1995

Phosphorylation of the human estrogen receptor by mitogen-activated protein kinase and casein kinase II: consequence on DNA binding.

We determined the amino acid and radiolabel sequences of tryptic [32P]phosphopeptides of the purified human estrogen receptor (hER) from MCF-7 cells and Sf9 cells. Serine 118 was identified as a site that was phosphorylated independently of estradiol-binding in MCF-7 cells. Proline is on the carboxy terminus of serine 118, which suggests that the serine-proline may be a consensus phosphorylation site motif for either the mitogen-activated protein (MAP) kinase or p34cdc2 kinase. MAP kinase selectively phosphorylated the recombinant hER in vitro on serine 118 independent of estradiol-binding, whereas p34cdc2 did not phosphorylate the hER. We demonstrated previously that serine 167 of the hER was phosphorylated in an estradiol-dependent manner. We therefore compared the consequence of hER phosphorylation at serine 118 by MAP kinase and phosphorylation at serine 167 by casein kinase II on the receptor's affinity for specific DNA binding. The binding of the hER to an estrogen response element was not altered by phosphorylation with MAP kinase at serine 118 but was significantly increased when phosphorylated at serine 167 by casein kinase II. These data suggest that phosphorylation of the hER by MAP kinase(s) pathways may influence receptor action by a mechanism other than the estradiol-dependent phosphorylation of hER by casein kinase II.

Adenosine Triphosphate↗
PubMed1990

Phosphorylation and activation of p40 tyrosine kinase by casein kinase-1.

Because examination of regulatory trans-phosphorylations can help elucidate the cellular functions of tyrosyl protein kinases, we have investigated the effects of phosphorylation by casein kinase-1 on the activity of the p40 tyrosyl protein kinase. We find that casein kinase-1 can phosphorylate the p40 tyrosyl kinase on serine and threonine residues, in part on a unique tryptic peptide. The phosphorylation induces a substantial increase in the tyrosyl protein kinase activity of p40, in contrast to most instances in which serine/threonine phosphorylation inhibits activity of tyrosyl protein kinases. These findings raise the possibility that p40 might be part of a protein phosphorylation network in which casein kinase-1 participates.

Adenosine Triphosphate↗
PubMed1992

Selective purification of two distinct protein kinases (C-kinase and casein kinase II) from the membrane fraction of mouse brain by NED-affinity column chromatography.

By means of NED-affinity column chromatography, two distinct protein kinases have been selectively purified from the crude membrane extract of mouse brain. One (designated P-I kinase) was eluted from the column by the buffer containing 5 mM EGTA and the other (designated P-II kinase) was eluted by the buffer containing 0.6 M KCl. The activity of A-kinase was detected in the column passed through fraction. Biochemical characteristics of P-I and P-II kinases corresponded exactly to those of C-kinase and casein kinase II (CK-II), respectively. In addition, immunoprecipitate experiment using anti-CK-II antiserum against the beta-subunit of Drosophila CK-II showed that P-II kinase is identical to CK-II and the 62 kDa cellular polypeptide is associated with the kinase.

Animals↗