Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.7.11.22 (cdc2)
 8,319 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We identified two major substrates for the proline-directed protein kinases--cdc2 kinase and tau protein kinase II (TPKII)--in the cytosol fraction from rat brains. The molecular masses of the proteins were 80 and 46 kDa. Because the 80-kDa protein was phosphorylated by protein kinase C and was heat stable, we examined the possibility that the protein might be myristoylated alanine-rich C kinase substrate (MARCKS). On the basis of a comparison between the properties of the 80-kDa protein and purified MARCKS, we concluded that the 80-kDa protein is indeed MARCKS. The amounts of phosphate incorporated into MARCKS by protein kinase C, cdc2 kinase, and TPKII were 1.7, 1.4, and 0.6 mol/mol of the protein, respectively. Two-dimensional tryptic peptide mapping indicated that phosphorylation sites by protein kinase C and proline-directed protein kinases completely differed. Only the seryl residue was phosphorylated by protein kinase C, whereas both seryl and threonyl residues were phosphorylated by cdc2 kinase and TPKII. Phosphorylation of MARCKS by protein kinase C inhibited the binding to calmodulin, whereas phosphorylation by cdc2 kinase and TPKII significantly increased the binding to calmodulin. The holoenzyme of protein phosphatase 2A dephosphorylated MARCKS that had been phosphorylated by protein kinase C, cdc2 kinase, or TPKII, whereas calcineurin was unable to dephosphorylate it. These results suggest that cdc2 kinase and TPKII regulate the functions of MARCKS in different ways from protein kinase C.
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PMID:Phosphorylation of myristoylated alanine-rich C kinase substrate (MARCKS) by proline-directed protein kinases and its dephosphorylation. 761 38

The activities of nuclear histone-H1 kinase and C-kinase as well as the amount of phosphate bound to histone-H1 following partial hepatectomy were studied in rat. It was found that the nuclear histone-H1 kinase activity increased twice within 80 h, first 20 to 30 h, and second at 50 to 70 h after partial hepatectomy. The timing of increase of the enzyme activity correlated with increased amount of bound phosphate. On the other hand, the increase of the C-kinase activities occurred between 5 and 15 h after partial hepatectomy. Antibodies raised against human cdk2, human cyclin-A and mouse cdc2 kinase showed no detectable effect on the nuclear histone H1 kinase activity. These results suggest that phosphorylation of histone-H1 in liver regeneration may be catalysed by a putative kinase(s).
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PMID:Role of nuclear histone-H1 kinase in regeneration of rat liver. 770 10

We identified the phosphorylation sites of glial fibrillary acidic protein (GFAP) for cdc2 kinase and Ca(2+)-calmodulin (CaM)-dependent protein kinase II. GFAP was phosphorylated to approximately 0.2 mol of phosphate/mol of GFAP by cdc2 kinase, and this phosphorylation did not induce disassembly of the filament structure. On the other hand, GFAP was phosphorylated to approximately 1.9 mol of phosphate/mol of GFAP by Ca(2+)-CaM-dependent protein kinase II, and this phosphorylation did induce disassembly of the filament. Sequential analysis of the purified phosphopeptides revealed that only Ser8 on GFAP was phosphorylated by cdc2 kinase, whereas Ser13, Ser17, Ser34, and Ser389 on GFAP were phosphorylated by Ca(2+)-CaM-dependent protein kinase II.
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PMID:Identification of phosphorylation sites on glial fibrillary acidic protein for cdc2 kinase and Ca(2+)-calmodulin-dependent protein kinase II. 782 64

cdc2-cyclin B activates protein phosphatase-1 (PP1) "in vitro", phosphorylates both catalytic subunit and inhibitor-2 (I2) and both processes are inhibited by a cdc2-inhibitory peptide. We compared the phosphorylation of I2 by cdc2-cyclin B and by the PP1-activator Glycogen Synthase Kinase 3 (GSK3). Each kinase introduced less than 0.1 mol phosphate/mol into I2 bound to PP1 and the same two tryptic phosphopeptides were obtained from I2, which contained phospho-T only. The same results were obtained also with isolated I2 phosphorylated by GSK3. Since GSK3 phosphorylates only T-72, cdc2-cyclin B is also likely to phosphorylate this site. This was confirmed by using I2 that had been mutated at this site. On the other hand cdc2-cyclin B introduced up to 0.8 mol/mol phosphate into isolated I2 and four phosphopeptides were obtained. The two new peptides contained phospho-T and one of them also phospho-S. These data indicate the presence of at least one T and one S that are phosphorylated only by cdc2-cyclin B and are accessible on isolated I2 only.
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PMID:Phosphorylation of the inhibitor-2 of protein phosphatase-1 by cdc2-cyclin B and GSK3. 786 66

A 100kD microtubule-bundling protein dynamin was phosphorylated in vitro by cdc2 kinase to approximately 1 mol of phosphate/mol of dynamin at a serine residue. These phosphorylations of dynamin greatly reduced its binding ability to microtubules.
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PMID:Phosphorylation of dynamin by cdc2 kinase. 804 26

1H-NMR and 31P-NMR spectroscopy were employed to assess the electrostatic consequences of phosphorylation of single and multiple tyrosine residues in peptides derived from the core and tail autophosphorylation regions of the human insulin receptor tyrosine-kinase domain. In both peptides, phosphorylation was accompanied by changes in the resonances from basic side-chains; those from acidic residues were unaffected. Tyrosine phosphorylation caused increases of up to one in the pKa values of histidine residues situated up to eight residues away in the primary sequence. Titration curve analysis by Hill plots suggested some cooperativity of histidine and phosphate ionizations. Behaviour closely analogous to that of the insulin receptor tail peptide was observed during changes in phosphorylation of the intact insulin receptor kinase domain, suggesting that the electrostatic dissemination effects seen for the isolated peptide are retained by the peptide sequence in the context of the much larger protein. Similar changes in the behaviour of basic residues were also observed upon tyrosine phosphorylation of a cdc2-derived peptide, suggesting that this potential of phosphorylation events to propagate directed structural changes may find a widespread utility in the activation of protein kinases and in the transduction of phosphorylation-based signalling.
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PMID:Phosphorylation effects on flanking charged residues. Structural implications for signal transduction in protein kinases. 807 32

We used the interaction trap, a yeast genetic selection for interacting proteins, to isolate human cyclin-dependent kinase interactor 1 (Cdi1). In yeast, Cdi1 interacts with cyclin-dependent kinases, including human Cdc2, Cdk2, and Cdk3, but not with Ckd4. In HeLa cells, Cdi1 is expressed at the G1 to S transition, and the protein forms stable complexes with Cdk2. Cdi1 bears weak sequence similarity to known tyrosine and dual specificity phosphatases. In vitro, Cdi1 removes phosphate from tyrosine residues in model substrates, but a mutant protein that bears a lesion in the putative active site cysteine does not. Overexpression of wild-type Cdi1 delays progression through the cell cycle in yeast and HeLa cells; delay is dependent on Cdi1 phosphatase activity. These experiments identify Cdi1 as a novel type of protein phosphatase that forms complexes with cyclin-dependent kinases.
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PMID:Cdi1, a human G1 and S phase protein phosphatase that associates with Cdk2. 824 50

Phosphorylation of the regulatory light chain of myosin II (MLC) controls the contractility of actomyosin in nonmuscle and muscle cells. It has been reported that cdc2 phosphorylates MLC in vitro at Ser-1 or Ser-2 and Thr-9 which protein kinase C phosphorylates (Satterwhite, L. L., M. J. Lohka, K. L. Wilson, T. Y. Scherson, L. K. Cisek, J. L. Corden, and T. D. Pollard. 1992 J. Cell Biol. 118:595-605). We have examined in vivo phosphorylation of MLC during mitosis and after the release of mitotic arrest. Phosphate incorporation of MLC in mitotic cells is found to be 6-12 times greater than that in nonmitotic cells. Phosphopeptide maps have revealed that the MLC from mitotic cells is phosphorylated at Ser-1 and/or Ser-2 (Ser-1/2), but not at Thr-9. MLC is also phosphorylated to a much lesser extent at Ser-19 which myosin light chain kinase phosphorylates. On the other hand, MLC of nonmitotic cells is phosphorylated at Ser-19 but not at Ser-1/2. The extent of phosphate incorporation is doubled at 30 min after the release of mitotic arrest when some cells start cytokinesis. Phosphopeptide analyses have revealed that the phosphorylation at Ser-19 is increased 20 times, while the phosphorylation at Ser-1/2 is decreased by half. This high extent of MLC phosphorylation at Ser-19 is maintained for another 30 min and gradually decreased to near the level of interphase cells as cells complete spreading at 180 min. On the other hand, phosphorylation at Ser-1/2 is decreased to 18% at 60 min, and is practically undetectable at 180 min after the release of mitotic arrest. The stoichiometry of MLC phosphorylation has been determined by quantitation of phosphorylated and unphosphorylated forms of MLC separated on 2D gels. The molar ratio of phosphorylated MLC to total MLC is found to be 0.16 +/- 0.06 and 0.31 +/- 0.05 in interphase and mitotic cells, respectively. The ratio is increased to 0.49 +/- 0.05 at 30 min after the release of mitotic arrest. These results suggest that the change in the phosphorylation site from Ser-1/2 to Ser-19 plays an important role in signaling cytokinesis.
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PMID:In vivo phosphorylation of regulatory light chain of myosin II during mitosis of cultured cells. 829 96

Human cyclins A and B1 were assembled with the cdk2 or cdc2 protein to reconstitute their respective kinase activities in vitro. Both cyclins complemented either cdk2 or cdc2, yielding kinase activities that supported the phosphorylation of histone H1. Activation of cdk2-catalyzed H1 kinase activity by cyclin A required a 10-min preincubation of the two components, whereas cdc2 kinase supported phosphate incorporation without a detectable time lag upon the addition of cyclin B1, suggesting a slower association rate of cdk2 with cyclin A compared with cdc2 and cyclin B1. Both cdk2 and cyclin A, as well as cdc2 and cyclin B1, formed stable complexes in the absence of ATP and substrate that could be isolated after glycerol gradient centrifugation. Incubation of the isolated complexes with ATP and histone H1 supported the phosphorylation of the substrate. Cyclin A-activated cdk2 or cdc2 phosphorylated p107, a pRB-related cellular protein, 10 times more effectively than the cyclin B1-complexed kinases. This was most likely due to a direct association of cyclin A with p107 (Ewen, M. E., Faha, B., Harlow, E., and Livingston, D. (1992) Science 255, 85-87; Faha, B., Ewen, M. E., Tsai, L.-H., Livingston, D., and Harlow, E. (1992) Science 255, 87-90). The reconstituted cdc2-cyclin B1 complex incorporated 4-5-fold more phosphate into the p34 subunit of the three-subunit (p70, p34, and p14) human single-stranded DNA-binding protein (also called RP-A), a DNA replication and DNA repair factor, than cdc2-cyclin A. No detectable phosphorylation of the p34 protein was observed with cdk2 complexed with either cyclin B1 or A. These data indicate that both cyclins as well as the catalytic subunits are important factors in controlling the rate of phosphorylation of a given substrate. The cyclin-activated cdc2 family kinases may target their cellular substrates through cyclin-mediated protein-protein interactions.
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PMID:Characterization of the in vitro reconstituted cyclin A or B1-dependent cdk2 and cdc2 kinase activities. 839 7

RNA polymerase II is a multisubunit enzyme composed of two large subunits of molecular weight in excess of 100,000 and a collection of 8-10 smaller subunits. The largest subunit, designated IIa, contains at its carboxyl terminus a highly repetitive domain consisting of tandem repeats of the consensus sequence Tyr-Ser-Pro-Thr-Ser-Pro-Ser. Extensive phosphorylation within this COOH-terminal domain (CTD) gives rise to subunit IIo which has a markedly reduced mobility in SDS-polyacrylamide gel electrophoresis (PAGE) relative to subunit IIa. Recent evidence suggests that RNA polymerase IIA, containing an unphosphorylated CTD, is involved in preinitiation complex assembly, whereas RNA polymerase IIO is involved in elongation. Consequently, CTD phosphorylation is thought to occur after RNA polymerase II has bound to the promoter by a protein kinase that stably associates with the preinitiation complex. We present here the partial purification and characterization of two distinct CTD kinases from a HeLa cell transcription extract. These CTD kinases, designated CTDK1 and CTDK2, are fractionated by chromatography on Mono Q. CTDK1 catalyzes the incorporation of approximately 33 pmol of phosphate/pmol of calf thymus RNA polymerase subunit IIa, almost exclusively on serine. CTDK2 catalyzes the incorporation of approximately 50 pmol of phosphate/pmol of calf thymus subunit IIa, predominantly on serine; appreciable phosphate transfer onto threonine is also observed. Phosphorylation by CTDK2, but not CTDK1, results in a complete mobility shift in SDS-PAGE of subunit IIa to the position of IIo. CTDK1 can utilize ATP, dATP, or GTP as phosphate donor, whereas CTDK2 can utilize only ATP or dATP. The apparent Km for ATP is 30 microM for CTDK1 and 60 microM for CTDK2. CTDK1 and CTDK2 also differ in their protein substrate specificity. CTDK1 phosphorylates casein whereas CTDK2 does not. Neither kinase phosphorylates phosvitin or histone H1 to an appreciable extent. CTDK1 and CTDK2 do not appear to be related to cdc2 kinases as determined by their inability to phosphorylate H1 and their failure to react with antibodies directed against the cdc2 kinase. These results establish that a partially fractionated HeLa transcription extract contains two distinct CTD kinases that differ in their nucleotide requirements and in their patterns of CTD phosphorylation.
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PMID:Partial purification and characterization of two distinct protein kinases that differentially phosphorylate the carboxyl-terminal domain of RNA polymerase subunit IIa. 841 77


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