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)

Casein kinase 2 (CK2) is a ubiquitous, multifunctional protein-seryl/threonyl kinase that has been implicated in cellular regulation. Synthetic peptides were patterned after three highly conserved regions in CK2: the N terminus (CK2-NT); the lysine-rich, kinase subdomain III segment (CK2-III) (nomenclature of Hanks et al. (Hanks, S. K., Quinn, A. M., and Hunter, T. (1988) Science 241, 42-52)); and a 10-residue segment located near kinase subdomain X that is shared between CK2 and p34cdc2 (CK2/cdc2). The CK2-III and CK2/cdc2 peptides markedly stimulated the autophosphorylation of the alpha- and alpha'-subunits of purified CK2 from sea star oocytes, and they elicited up to 2-fold increases in its casein or phosvitin phosphotransferase activity. These peptides completely reversed nearly total inhibition of CK2 phosphotransferase activity toward itself, casein, and phosvitin by either heparin or poly(Glu,Tyr; 4:1), whereas CK2-NT was ineffective. Elution of CK2 from heparin-agarose with the CK2-III peptide indicated that this region of CK2 might mediate heparin binding to CK2. Affinity-purified rabbit polyclonal antibodies developed against both CK2-III and CK2/cdc2, but not CK2-NT, also produced up to 1.8-fold enhancements of the casein and phosvitin phosphotransferase activities of purified CK2. All three of the antipeptide antibody preparations immunoreacted with the alpha- and alpha'-subunits of CK2 on Western blots. These studies indicate that kinase subdomains III and X are involved in the modulation of CK2 phosphotransferase activity.
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PMID:Structure-function analysis of casein kinase 2 with synthetic peptides and anti-peptide antibodies. 131 43

Human cyclin B1-bound cdc2 kinase phosphorylated the threonine residue in the sequence -Thr-Pro-Lys-Lys-Ala- but hardly phosphorylated it in the sequence -Thr-Pro-Lys-Ala-Lys. The sequence -Thr-Pro-Ala-Pro-Lys-, as found in p53 protein, was also phosphorylated by this enzyme, but less efficiently than in the sequence described above. When the threonine residue in -Thr-Pro-Lys-Lys-Ala- was changed to a serine or a tyrosine residue, the enzyme phosphorylated the serine, but not the tyrosine residue. Changing the lysine next to the proline to alanine reduced its efficiency as a substrate. The peptide, Ala-Ala-Ala-Ala-Lys-Thr-Pro-Ala-Lys-Ala-Ala, containing the -Thr-Pro-Ala-Lys- sequence, but not the other lysine residues, was not used as a substrate by the kinase.
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PMID:Preference of human cdc2 kinase for peptide substrate. 145 May 22

Neurofilament (NF) protein [high molecular mass (NF-H)] is extensively phosphorylated in vivo. The phosphorylation occurs mainly in its characteristic KSP (Lys-Ser-Pro) repeat motifs. There are two major types of KSP motifs in the NF-H tail domain: KSPXKX and KSPXXX. Recent studies by two different laboratories have demonstrated the presence of a cdc2-like kinase [cyclin-dependent kinase-5 (cdk5)] in nervous tissue that selectively phosphorylates KSPXKX and XS/TXK motifs in NF-H and lysine-rich histone (H1). This article describes the identification of phosphatases dephosphorylating three different substrates: histone (H1), NF-H in a NF preparation, and a bacterially expressed C-terminal tail domain of NF-H, each containing KSPXKX repeats phosphorylated in vitro by cdk5. Among various phosphatases identified, protein phosphatase (PP) 2A from rabbit skeletal muscle appeared to be the most effective phosphatase in in vitro assays. Three phosphatase activity peaks--P1, P2, and P3--were partially purified from frozen rat spinal cord by ion exchange and size exclusion column chromatography and then characterized on the basis of biochemical, pharmacological, and immunochemical studies. One of the three peaks was identified as PP2A, whereas the others were mixtures of both PP2A and PP1. These three peaks could dephosphorylate cdk5-phosphorylated 32P-histone (H1), 32P-NF-H in the NF preparation, and 32P-NF-H tail fusion protein. These studies suggest the involvement of PP2A or a PP2A-like activity in the regulation of the phosphorylation state of KSPXKX motifs in NF-H.
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PMID:Neuronal cyclin-dependent kinase-5 phosphorylation sites in neurofilament protein (NF-H) are dephosphorylated by protein phosphatase 2A. 776 48

Synthetic peptide representing the site Ser-41 in vimentin, Leu-Gly-Ser41-Ala42-Leu-Arg44-Arg-Arg-NH2, and its analogs in which Ala-42 was replaced by various amino acids were tested as substrates for cdc2 kinase. Among them, the analog containing sarcosine as well as proline was an excellent substrate. The result suggests that the N-substituted structure of proline immediately following the site is important for cdc2 kinase phosphorylation. Replacement of Ala-42 by polar amino acids, especially lysine, had negative effects on peptide phosphorylation. The peptides in this study were also assayed with another type of proline-directed protein kinase, tau protein kinase II. The substrate specificity differed essentially from that of cdc2 kinase.
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PMID:Phosphorylation of synthetic vimentin peptides by cdc2 kinase. 837 19

We have studied the influence of a temperature-sensitive cdc2-1 mutation in DNA polymerase delta on the selection-induced mutation occurring at the LYS-2 locus in the yeast Saccharomyces cerevisiae. It was found that in cells plated on synthetic complete medium lacking only lysine, the numbers of Lys+ revertant colonies accumulated in a time-dependent manner in the absence of any detectable increase in cell number. When cdc2-1 mutant cells, after selective plating, were incubated at the restrictive temperature of 37 degrees C for 5 h daily for 7 days, the frequency of an adaptive reversion of lys(-)-->Lys+ was significantly higher than the frequency in cells incubated only at the permissive temperature, or in wild-type cells incubated either at 23 degrees C or 37 degrees C. Therefore, when the proofreading activity of DNA polymerase delta is impaired under restrictive conditions, the frequency of adaptive mutations is markedly enhanced.
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PMID:Effects of the CDC2 gene on adaptive mutation in the yeast Saccharomyces cerevisiae. 859 82

The mammalian p34cdc2 protein kinase, a universal cell cycle regulator, complements cdc2/CDC28 temperature-sensitive mutations in yeasts. We report the biochemical characterisation of two substitutions of human cdc2 at lysine 33, a residue involved in nucleotide binding, that differently alter the fission yeast cell cycle. K33A-hscdc2 and K33R-hscdc2 mutants are both catalytically inactive, but overexpression of K33R-cdc2 is lethal while K33A-cdc2 is not. We show that human K33R-cdc2 acts as a dominant negative allele that associates yeast cdc13/cyclinB and therefore renders endogeneous Schizosaccharomyces pombe cdc2 unactivatable. These results are discussed on the light of the molecular modeling of the mutants in the cdc2 model structure.
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PMID:Characterisation of human cdc2 lysine 33 mutations expressed in the fission yeast Schizosaccharomyces pombe. 860 92

Neuronal cdk5 can phosphorylate certain lys-ser-pro (KSP) motifs of neurofilaments and tau protein in the nervous system. We have immunoprecipitated the cdk5 from rat brain using a polyclonal antibody raised against the C-terminus of cdk5. The immunoprecipitate has phosphorylated a KSPXK peptide analog of NF-H, as well as histone H1 and a bacterially expressed rat NF-H protein. The kinase activity was inhibited by staurosporine, isopentanyladenine and olomoucine in a dose dependent manner. Kinetic studies indicated Ki values of 39 nM, 38 microM and 8 microM, respectively for staurosporine, isopentanyladenine and olomoucine. The inhibition by staurosporine was non-competitive with respect to phosphoryl acceptor acceptor substrates. Western blot analysis of the immunoprecipitate showed both cdk5 and p67 (Munc-18), a putative regulator molecule of the kinase. Addition of p67 fusion protein enhanced the kinase activity of the immunoprecipitate by 60% above the basal activity. P67 elevated Ki values for both staurosporine and olomoucine. The degree of inhibition at high concentrations of these inhibitors was unaltered by exogenous p67 indicating a lack of competitive interactions with p67. The high affinity of staurosporine for cdk5 suggests that cdk5 may be one of the targets for the neurotropic effect of staurosporine.
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PMID:Inhibition of neuronal cyclin-dependent kinase-5 by staurosporine and purine analogs is independent of activation by Munc-18. 872 73

Immediately before the transition from metaphase to anaphase, the protein kinase activity of maturation or M-phase promoting factor (MPF) is inactivated by a mechanism that involves the degradation of its regulatory subunit, cyclin B. The availability of biologically active goldfish cyclin B produced in Escherichia coli and purified goldfish proteasomes (a nonlysosomal large protease) has allowed the role of proteasomes in the regulation of cyclin degradation to be examined for the first time. The 26S, but not the 20S proteasome, digested recombinant 49-kD cyclin B at lysine 57 (K57), producing a 42-kD truncated form. The 42-kD cyclin was also produced by the digestion of native cyclin B forming a complex with cdc2, a catalytic subunit of MPF, and a fragment transiently appeared during cyclin degradation when eggs were released from metaphase II arrest by egg activation. Mutant cyclin at K57 was resistant to both digestion by the 26S proteasome and degradation at metaphase/anaphase transition in Xenopus egg extracts. The results of this study indicate that the destruction of cyclin B is initiated by the ATP-dependent and ubiquitin-independent proteolytic activity of 26S proteasome through the first cutting in the NH2 terminus of cyclin (at K57 in the case of goldfish cyclin B). We also surmise that this cut allows the cyclin to be ubiquitinated for further destruction by ubiquitin-dependent activity of the 26S proteasome that leads to MPF inactivation.
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PMID:Initiation of cyclin B degradation by the 26S proteasome upon egg activation. 929 86

Neurofilaments (NFs), the neuron-specific intermediate (i.e. approximately 10-nm diameter) filaments are major cytoskeletal components of most neurons. In a mature mammalian neuron, NFs are co-assembled from three subunits, NF-L (low), NF-M (medium), and NF-H (high), with molecular masses of 68, 95, and 115 kDa, respectively. Neurofilament proteins (NF-Ps), particularly, NF-H, are most extensively phosphorylated in large myelinated axons under normal conditions. This phosphorylation occurs on the serine residues of the lysine (Lys)-serine (Ser)-proline (Pro) (KSP) multiple amino acid repeats of the carboxy-terminal tail domain. Phosphorylation of KSP motifs affects physical, biochemical, and immunological properties of NF-H. For example, phosphorylation is thought to play a pivotal role in the maintenance of the neuronal cytoskeletal structure which influences the conduction velocity of the nerve fiber. The key components responsible for phosphorylation are not known. In this study, an identified cyclin-dependent kinase 5 (cdk5), isolated from nervous tissue, has been shown to phosphorylate the human NF-H (hNF-H) and affects its electrophoretic mobility. On the basis of the following observations, it is suggested that neuronal cdk5 (cdk5) phosphorylates KSPXK motifs in the human high molecular weight neurofilament (hNF-H) and affects its electrophoretic mobility. (1) A 14-mer synthetic peptide (KSPEKAKSPVKEEA) derived from hNF-H; (2) a bacterially expressed protein containing 14 KSPXK multiple repeats of hNF-H in C-terminal tail domain; and (3) a dephosphorylated hNF-H in neurofilament preparation are phosphorylated by cdk5. The decrease in molecular mass of hNF-H caused by dephosphorylated was completely recovered upon cdk5 phosphorylation. It is proposed that neuronal cdk5 regulates phosphorylation of the KSPXK motif in hNF-H and other cytoskeletal proteins with similar motifs in the nervous system.
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PMID:Phosphorylation of human high molecular weight neurofilament protein (hNF-H) by neuronal cyclin-dependent kinase 5 (cdk5). 931 98

We studied spheroid (multicellular aggregate) formation by hepatocytes and the expression of liver-specific functions such as albumin secretion when hepatocytes were cultured with various extracellular matrices. Hepatocytes cultured on Primaria(R) and poly-D-lysine coated dishes, and in the presence of a polymer, Eudragit, formed spheroids, and they also exhibited higher liver-specific functions and poor growth compared to monolayer cultures. The results indicated that the cell morphological change and cell-cell interaction caused by the spheroid formation were key factors promoting the expression of the liver-specific functions. To elucidate the mechanism underlying the poor growth in spheroids, we examined the HGF signaling pathway. Phosphorylation and down-regulation of the HGF receptor (c-Met proto-oncogene product) were observed for the cells from both monolayer and spheroid cultures, but Ras activation was partly blocked in spheroids. Furthermore, we found that CDK inhibitors, p21 and p27, were highly expressed in spheroids. These results suggested that the reduced Ras signaling and high expression of the CDK inhibitors might cause the lower growth in spheroids. We then examined the relationship between liver-enriched transcription factors (C/EBPalpha and beta) and liver-specific functions. The results revealed that the high expression of C/EBPalpha was maintained during cultures when hepatocytes formed spheroids. Antisense oligonucleotides of C/EBPalpha repressed albumin secretion and the expression of p21, suggesting that the transcription factor, C/EBPalpha, may play a crucial role in the growth and differentiation of hepatocytes in spheroids.
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PMID:Differentiation and proliferation of primary rat hepatocytes cultured as spheroids. 979 21


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