Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:2.7.11.1 (protein kinase)
81,284 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A new protein kinase has been characterized among the proteins tightly bound to rat liver DNA and released by DNase I and RNase A treatment. This enzyme was separated by gel filtration from this released material. Its apparent molecular mass was found to be 34 kDa and it is made of a single unit. The main characteristic of this protein kinase is that it is arginine-specific. Isolation of phosphoarginine required the use of proteolytic enzymes at alkaline pH since the phosphate bond is highly acid-labile. This protein kinase is able to autophosphorylate and to phosphorylate a single chromosomal protein of 11 kDa also tightly bound to DNA. It uses ATP and dATP as phosphate donors and is cAMP-independent. Its optimal activity requires Mn2+ ions. Vanadate, spermine and heparin have no effect on its activity.
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PMID:Characterization of an arginine-specific protein kinase tightly bound to rat liver DNA. 360 29

The kinetics of the interaction of the DNA double-helix-destabilizing protein from roe-deer liver with different DNAs revealed a fast phase which is observed both by the increase in A260 of the DNA and the quenching of the protein intrinsic fluorescence. A slow phase with a smaller amplitude is only recorded by the increase of A260.--The protein contains slightly less than two phosphate groups per molecule, removal of one of which by alkaline phosphatase does not affect its activity; however, removal of both phosphates decreases the DNA-unwinding property significantly. A similar decrease in activity is also revealed upon incorporation of an additional phosphate by cAMP-dependent protein kinase I.--Results of the protection of poly[d(A--T)] from DNase I digestion by the protein are in favor of a migration of the protein along the DNA.
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PMID:The kinetics of the interaction of a helix-destabilizing protein from roe-deer liver with DNA and the influence of phosphorylation. 625 Sep 67

Nuclear envelopes were prepared from purified rat liver nuclei by lysis with heparin, digestion with deoxyribonuclease I (DNase I), or sonication. The envelopes were fractionated by centrifugation on sucrose density gradients and analyzed for protein kinase activity using endogenous and exogenous protein substrates and [gamma-32 P]ATP. The protein kinase activity toward endogenous proteins was markedly affected by the method used to isolate the envelopes, with sonication producing a preparation with very low activity. At least 12 phosphoproteins in nuclear envelopes isolated by the heparin or DNase I method were detected by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and autoradiography. A 32P-labeled material migrating with an apparent Mr = 3000 was extracted with chloroform:methanol:HCl and was identified as a mixture of phospholipids. Total 32P incorporation into nuclear envelopes peaked at 5 min of incubation, followed by a decrease in labeled products. This decrease was due to both phosphoprotein phosphatase activity and degradation of the lipid products. The highest protein kinase activity toward endogenous proteins was expressed with [gamma-32P]ATP in the presence of MgCl2; however, some phosphorylation also occurred with MnCl2, CoCl2, NiCl2, and [gamma-32P]GTP in the presence of MgCl2. Nuclear envelope protein phosphorylation was unaffected by cyclic nucleotides and calmodulin, slightly inhibited by CaCl2, MnCl2, CoCl2, disulfides, and sulfhydryl alkylating agents, and strongly inhibited by LaCl3 and phosphatidylglycerol. Nuclear porelamina complexes isolated from phosphorylated envelopes contained phosphoproteins of 7, 20, 51, 59, and 70 kDa. Incubation of pore-lamina complexes isolated from unlabeled envelopes with [gamma-32P]ATP resulted in 32P incorporation into the 20-, 51-, and 50-kDa proteins.
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PMID:Phosphorylation of rat liver nuclear envelopes. I. Characterization of in vitro protein phosphorylation. 630 4

A protein kinase has been characterized among the proteins tightly bound to DNA. It is not extracted with 1 M NaCl and is released by extensive DNase I digestion. This enzyme is able to phosphorylate nucleosomal histones, essentially H2B and H3, and several non-histone proteins associated with DNA, on serine residue(s). It does not phosphorylate protamine, casein, phosvitin and the chromosomal non-histone proteins extracted with 1 M NaCl and is cAMP independent. This protein kinase can be distinguished from the previously described enzymes.
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PMID:A new cAMP independent protein kinase tightly bound to DNA, in rat liver nuclei. 631 66

The rat tyrosine aminotransferase gene is a model system to study transcriptional regulation by glucocorticoid hormones. We analyzed transcription factor binding to the tyrosine aminotransferase gene glucocorticoid-responsive unit (GRU) at kb -2.5, using in vivo footprinting studies with both dimethyl sulfate and DNase I. At this GRU, glucocorticoid activation triggers a disruption of the nucleosomal structure. We show here that various regulatory pathways affect transcription factor binding to this GRU. The binding differs in two closely related glucocorticoid-responsive hepatoma cell lines. In line H4II, glucocorticoid induction promotes the recruitment of hepatocyte nuclear factor 3 (HNF3), presumably through the nucleosomal disruption. However, the footprint of the glucocorticoid receptor (GR) is not visible, even though a regular but transient interaction of the GR is necessary to maintain HNF3 binding. In contrast, in line FTO2B, HNF3 binds to the GRU in the absence of glucocorticoids and nucleosomal disruption, showing that a "closed" chromatin conformation does not repress the binding of certain transcription factors in a uniform manner. In FTO2B cells, the footprint of the GR is detectable, but this requires the activation of protein kinase A. In addition, protein kinase A stimulation also improves the recruitment of HNF3 independently of glucocorticoids and enhances the glucocorticoid response mediated by this GRU in an HNF3-dependent manner. In conclusion, the differences in the behavior of this regulatory sequence in the two cell lines show that various regulatory pathways are integrated at this GRU through modulation of interrelated events: transcription factor binding to DNA and nucleosomal disruption.
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PMID:Glucocorticoids and protein kinase A coordinately modulate transcription factor recruitment at a glucocorticoid-responsive unit. 756 84

Insulin-like growth factor I (IGF-I) is a locally synthesized anabolic growth factor for bone. IGF-I synthesis by primary fetal rat osteoblasts (Ob) is stimulated by agents that increase the intracellular cAMP concentration, including prostaglandin E2 (PGE2). Previous studies with Ob cultures demonstrated that PGE2 enhanced IGF-I transcription through selective use of IGF-I promoter 1, with little effect on IGF-I messenger RNA half-life. Transient transfection of Ob cultures with an array of promoter 1-luciferase reporter fusion constructs has now allowed localization of a potential cis-acting promoter element(s) responsible for cAMP-stimulated gene expression to the 5'-untranslated region (5'-UTR) of IGF-I exon 1, within a segment lacking a consensus cAMP response element. Our evidence derives from three principal observations: 1) a transfection construct containing only 122 nucleotides (nt) of promoter 1 and 328 nt of the 5'-UTR retained full PGE2-stimulated reporter expression; 2) maximal PGE2-driven reporter expression required the presence of nt 196 to 328 of exon 1 when tested within the context of IGF-I promoter 1; 3) cotransfection of IGF-I promoter-luciferase-reporter constructs with a plasmid encoding the alpha-isoform of the catalytic subunit of murine cAMP-dependent protein kinase (PKA) produced results comparable to those seen with PGE2 treatment, whereas cotransfection with a plasmid encoding a mutant regulatory subunit of PKA that cannot bind cAMP blocked PGE2-induced reporter expression. Deoxyribonuclease I footprinting of the 5'-UTR of exon 1 demonstrated protected sequences at HS3A, HS3B, and HS3D, three of six DNA-protein binding sites previously characterized with rat liver nuclear extracts. Of these three regions, only the HS3D binding site is located within the functionally identified hormonally responsive segment of IGF-I exon 1. These results directly implicate PKA in the control of IGF-I gene transcription by PGE2 and identify a segment of IGF-I exon 1 as being essential for this hormonal regulation.
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PMID:Regulation of insulin-like growth factor I transcription by cyclic adenosine 3',5'-monophosphate (cAMP) in fetal rat bone cells through an element within exon 1: protein kinase A-dependent control without a consensus AMP response element. 764 98

In hepatocytes, insulin-like growth factor-binding protein-1 (IGFBP-1) levels are increased by glucocorticoids and by agents that raise intracellular cAMP levels such as glucagon, theophylline, forskolin, and cAMP analogues. In contrast, insulin lowers IGFBP-1 levels, an effect dominant over the glucocorticoid and cAMP effects. Previous studies showed that dibutyryl cAMP (Bt2cAMP) and theophylline increase IGFBP-1 promoter activity in HEP G2 human hepatoma cells and that insulin abolishes this increase. In studies reported here, HEP G2 cells were used to further evaluate the role of cAMP in stimulating IGFBP-1 expression. Initial studies found that either 0.5 or 5.0 mM Bt2cAMP alone, or the combination of 0.5 mM Bt2cAMP and 2 mM theophylline, increased IGFBP-1 protein levels, mRNA levels, and promoter activity, but that the addition of theophylline to Bt2cAMP was required to give a approximately 5-fold increase in promoter activity. Deletion mutations of the IGFBP-1 promoter were used to show that much of the effect of Bt2cAMP and theophylline was conferred by the region between 269 and 246 base pairs (bp) 5' of the IGFBP-1 mRNA cap site. DNase I protection assays showed that HEP G2 nuclear extract footprinted the region from 273 to 249 bp 5' of the cap site; this region, designated P2, has a central CGTCA motif common to cAMP-responsive elements (CREs). Mutating the CGTCA motif in the 1205-bp IGFBP-1 promoter construct to TAGCA led to a 51% decrease in the ability of Bt2cAMP and theophylline to stimulate IGFBP-1 promoter activity above control levels. In addition, cotransfection of the catalytic subunit of cAMP-dependent protein kinase A (PKA) with the native 1205-bp IGFBP-1 promoter construct stimulated IGFBP-1 promoter activity 3.9-fold, but the TAGCA mutation decreased by 73% the ability of PKA to stimulate IGFBP-1 promoter activity above control levels. Mutating the CGTCA motif to TAGCA also blocked the ability of both crude HEP G2 nuclear extract and recombinant CRE-binding protein to bind to the P2 element. These data suggest that the P2 element is a CRE that confers at least part of the stimulatory effect of cAMP on the human IGFBP-1 promoter.
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PMID:Identification of a promoter element which participates in cAMP-stimulated expression of human insulin-like growth factor-binding protein-1. 768 58

Myotonic dystrophy is caused by an expansion of a CTG triplet repeat sequence in the 3' noncoding region of a protein kinase gene, yet the mechanism by which the triplet repeat expansion causes disease remains unknown. This report demonstrates that a DNase I hypersensitive site is positioned 3' of the triplet repeat in the wild-type allele in both fibroblasts and skeletal muscle cells. In three unrelated individuals with myotonic dystrophy that have large expansions of the triplet repeat, the allele with the triplet repeat expansion exhibited both overall DNase I resistance and inaccessibility of nucleases to the adjacent hypersensitive site. These results indicate that the triplet repeat expansion alters the adjacent chromatin structure, establishing a region of condensed chromatin, and suggests a molecular mechanism for myotonic dystrophy.
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PMID:Triplet repeat expansion in myotonic dystrophy alters the adjacent chromatin structure. 777 32

To analyze regulation of the human T-cell leukemia virus type I (HTLV-I) long terminal repeat (LTR), cell lines were generated from LTR-tax x LTR-beta-galactosidase (beta-Gal) doubly transgenic mouse fibroblastic tumors. The HTLV-I LTR directs expression of both the tax and lacZ genes, and Tax up-modulates both promoters in primary cells. However, once cells were transformed by tax, beta-Gal but not tax expression was suppressed. Supertransformation of these cells with v-src suppressed both beta-Gal and tax expression. This suppression was reversed by treatment with the tyrosine kinase inhibitor herbimycin A or protein kinase A inhibitor H8. Electrophoretic mobility shift assays demonstrated augmented binding in the R but not U3 region. This binding was competitively inhibited by a high-affinity CREB oligodeoxynucleotide and super-shifted with a specific CREB antibody. Treatment of cells with the cyclic AMP analog dibutyryl cyclic AMP also transiently increased the R region binding dramatically. In vitro DNase I footprint analysis identified a protein-binding sequence in the R region which corresponded with suppression. However, this target sequence lacked a conventional CREB-binding site. A 70.5-kDa DNA-binding protein was partially purified by affinity chromatography, along with a 49-kDa protein which reacted with CREB-specific sera. These data demonstrate that HTLV-I LTR suppression is associated with CREB factor binding in the R region, probably by direct interaction with a 70.5-kDa protein, and provide a novel mechanism for maintenance of viral latency.
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PMID:Transcriptional suppression of the human T-cell leukemia virus type I long terminal repeat occurs by an unconventional interaction of a CREB factor with the R region. 803 15

Casein kinase 2 was released from rat liver cells nuclei by digestion with DNase I plus RNase A. This treatment also released three major substrates of 50, 40-42, and 37 kDa. Casein kinase 2 and substrates were also extracted by DNase or RNase separately. However, in DNase extracts only the 37 kDa protein was phosphorylated by casein kinase 2, whereas in RNase extracts all three substrates were phosphorylated. When the DNase extracts were subsequently treated with RNase the 40-42 substrates were then phosphorylated, indicating that their interaction with RNA prevents their phosphorylation by casein kinase 2. The ratio of B: alpha subunits of casein kinase 2 present in the nuclease extracts was higher than that of the purified enzyme, which is assumed to be 1:1. A further analysis by sucrose gradient centrifugation revealed that under physiological salt conditions casein kinase 2 from nuclease extracts formed large aggregates (higher than 300 kDa) which were disrupted at 400 mM KCl. At the latter KCl concentration CK-2 activity was localized at a position corresponding to a M(r) of 230-250 kDa, which is still higher than the typical tetrameric form of the enzyme.
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PMID:Casein kinase 2 and protein substrates are released from rat liver cells nuclei by DNase or RNase digestion. 804 72


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