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)

Protein phosphatase 2A2 is inactivated by phosphorylation following incubation with purified preparations of an autophosphorylation-activated protein kinase (Hong Guo and Zahi Damuni (1992) Proc. Natl. Acad. Sci. U.S.A. 90, 2500-2504). This protein kinase was purified about 250,000-fold from extracts of bovine kidney to apparent homogeneity. The purified preparations exhibited a single polypeptide of apparent M(r) approximately 36,000. Up to 1 mol of phosphoryl groups was incorporated per mol of the purified kinase following incubation with ATP. This autophosphorylation reaction (t1/2 approximately 0.5-1 min) was accompanied by a approximately 10-fold activation of the kinase. Autophosphorylation and activation were reversed by protein phosphatase 2A2 or the catalytic subunit of protein phosphatase 1. Phosphoamino acid analysis indicated that the kinase underwent autophosphorylation on threonines. The rate of autophosphorylation was independent of the concentration of the enzyme and a slope of 0.97 (gamma = 0.998) was obtained by van't Hoff's plot indicating that autoposphorylation was intramolecular. Relative to myelin basic protein, the enzyme exhibited about 8, 62, 130, 33, 5, and < 0.1% activity with histones H1, H2A, H2B, H3, and H4 and with glycogen synthase alpha, respectively. Heparin inhibited the activity of the enzyme half-maximally at about 20 micrograms/ml. The results indicate that this autophosphorylation-activated kinase is a new protein kinase.
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PMID:Purification and characterization of an autophosphorylation-activated protein serine threonine kinase that phosphorylates and inactivates protein phosphatase 2A. 838 87

The effects of UV light or fungal elicitors on plant cells have so far been studied mostly with respect to defense-related gene activation. Here, an inverse correlation of these stimulatory effects with the activities of several cell cycle-related genes is demonstrated. Concomitant with the induction of flavonoid biosynthetic enzymes in UV-irradiated cell suspension cultures of parsley (Petroselinum crispum), total histone synthesis declined to about half the initial rate. A subclass of the histone H3 gene family was selected to demonstrate the close correlation of its expression with cell division, both in intact plants and cultured cells. Using RNA-blot and run-on transcription assays, it was shown that one arbitrarily selected subclass of each of the histone H2A, H2B, H3 and H4 gene families and of the genes encoding a p34cdc2 protein kinase and a mitotic cyclin were transcriptionally repressed in UV-irradiated as well as fungal elicitor-treated parsley cells. The timing and extent of repression differed between the two stimuli; the response to light was more transient and smaller in magnitude. These differential responses to light and elicitor were inversely correlated with the induction of phenylalanine ammonia-lyase, a key enzyme of phenylpropanoid metabolism. Essentially the same result was obtained with a defined oligopeptide elicitor, indicating that the same signaling pathway is responsible for defense-related gene activation and cell cycle-related gene repression. A temporary (UV light) or long-lasting (fungal elicitor) cessation of cell culture growth is most likely due to an arrest of cell division which may be a prerequisite for full commitment of the cells to transcriptional activation of full commitment of the cells to transcriptional activation of pathways involved in UV protection or pathogen defense. This conclusion is corroborated by the observation that the histone H3 mRNA level greatly declined around fungal infection sites in young parsley leaves.
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PMID:Gene activation by UV light, fungal elicitor or fungal infection in Petroselinum crispum is correlated with repression of cell cycle-related genes. 858 Sep 59

The transcription factors Oct-1 and Oct-2 bind differentially to three octamer binding sequences corresponding to the octamer binding site from the H2B promoter [ATGCTAATAA], a simple TAATGARAT motif, found in herpes simplex virus IE4/5 genes [GCGGTAATGAGAT], and a perfect consensus overlapping octamer/TAATGARAT motif [ATGCTAATGAGAT]. By comparing the effects of protein kinase A, protein kinase C and casein kinase 2 in vitro on the binding of Oct-1 and Oct-2 to the three motifs, we show that the actions of these kinases regulate Oct-1 and Oct-2 DNA binding independently of each other in a binding-site-specific manner. Inhibition of cellular phosphatases also regulate Oct-1 and Oct-2 DNA binding in a binding-site-specific manner. Both kinase and phosphatase activity are important for regulating the DNA binding activity of Oct-1 and Oct-2 because, in the presence of phosphatase inhibitors, protein kinase A attenuates the binding of both Oct-1 and Oct-2 to the octamer binding site but enhances binding when phosphatase inhibitors are omitted. Thus the DNA specificity of Oct-1 and Oct-2 can be regulated in vitro by the action of different kinases.
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PMID:Oct-1 [corrected] and Oct-2 DNA-binding site specificity is regulated in vitro by different kinases. 864 73

Transcription of the genes for the human histone proteins H4, H3, H2A, H2B, and H1 is activated at the G1/S phase transition of the cell cycle. We have previously shown that the promoter complex HiNF-D, which interacts with cell cycle control elements in multiple histone genes, contains the key cell cycle factors cyclin A, CDC2, and a retinoblastoma (pRB) protein-related protein. However, an intrinsic DNA-binding subunit for HiNF-D was not identified. Many genes that are up-regulated at the G1/S phase boundary are controlled by E2F, a transcription factor that associates with cyclin-, cyclin-dependent kinase-, and pRB-related proteins. Using gel-shift immunoassays, DNase I protection, and oligonucleotide competition analyses, we show that the homeodomain protein CDP/cut, not E2F, is the DNA-binding subunit of the HiNF-D complex. The HiNF-D (CDP/cut) complex with the H4 promoter is immunoreactive with antibodies against CDP/cut and pRB but not p107, whereas the CDP/cut complex with a nonhistone promoter (gp91-phox) reacts only with CDP and p107 antibodies. Thus, CDP/cut complexes at different gene promoters can associate with distinct pRB-related proteins. Transient coexpression assays show that CDP/cut modulates H4 promoter activity via the HiNF-D-binding site. Hence, DNA replication-dependent histone H4 genes are regulated by an E2F-independent mechanism involving a complex of CDP/cut with cyclin A/CDC2/ RB-related proteins.
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PMID:CDP/cut is the DNA-binding subunit of histone gene transcription factor HiNF-D: a mechanism for gene regulation at the G1/S phase cell cycle transition point independent of transcription factor E2F. 887 67

A cAMP-dependent histone kinase was purified and characterized from spermatozoa of the sea urchin Hemicentrotus pulcherrimus. The molecular mass of the kinase was estimated to be 178 kDa by native PAGE and 400 kDa by gel chromatography on a Superose 6 HR 10/30 column. The enzyme, composed of two 39-kDa catalytic subunits and two 48-kDa regulatory subunits, phosphorylates the lysine-rich histone subspecies (H1 and H2B) isolated from H. pulcherrimus spermatozoa. We isolated cDNA clones encoding a 39-kDa catalytic subunit and a 48-kDa regulatory subunit of the enzyme. The cDNA clone for the 39-kDa subunit was 3881 bp, and the 352-residue deduced amino acid sequence showed 78% similarity with the catalytic subunit of/mammalian cAMP-dependent protein kinase (PKA). The cDNA for the 48-kDa subunit was 4589 bp and the 368-residue deduced amino acid sequence showed 57% similarity with the regulatory subunit of mammalian PKA, although the N-terminal 77 residues showed poor similarity. The mRNAs encoding both the catalytic subunit (7.5 kb) and the regulatory subunit (4.6 kb) were expressed in testis, ovary and egg. An inter-phylum hybrid enzyme, reconstituted from the regulatory subunit of cAMP-dependent histone kinase of sea urchin sperm and the catalytic subunit of bovine heart PKA, has a cAMP-dependent histone kinase activity. Thus, we suggest that the N-terminal 77-amino-acid residues of the regulatory subunit are not essential for inhibition by the regulatory subunit of the catalytic subunit, and that cAMP-dependent inhibitory activity of the regulatory subunit resides in the sequence between the inhibitory site and the C-terminus.
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PMID:Cyclic-AMP-dependent activation of an inter-phylum hybrid histone-kinase complex reconstituted from sea urchin sperm-regulatory subunits and bovine heart catalytic subunits. 905 23

Prothymosin alpha (ProTalpha) is an acidic protein involved in cell proliferation. Its phosphorylation status is correlated with proliferative activity. Here we report the isolation and characterization of a ProTalpha-phosphorylating kinase (ProTalphaK) from mouse splenocytes that seems to be responsible for the in vivo phosphorylation of ProTalpha and that differs from other protein kinases reported to date. This enzyme, mainly located in the cytosol, has an molecular mass of 180 kDa and appears to be made up of two proteins of 64 and 60 kDa. Its activity was markedly enhanced by mitogenic activation of cells. The ProTalpha residues phosphorylated by the enzyme in vitro are a Thr at position 7 and another Thr at positions 12 or 13, both located within casein kinase 2 (CK-2) consensus motifs; these are the same residues as are phosphorylated in vivo. The new enzyme shows a number of clear structural and catalytic differences from CK-2. It phosphorylates histones H2B and H3, although with weaker activity than ProTalpha. An enzyme with the same characteristics was also found in other murine tissues and cell lines.
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PMID:A 180-kDa protein kinase seems to be responsible for the phosphorylation of prothymosin alpha observed in proliferating cells. 909 94

A 74-kDa delta/B" subunit was isolated by heparin-Sepharose column chromatography from human erythrocyte protein phosphatase 2A (PP2A) consisting of a 34-kDa catalytic subunit (alpha/C) and 63- and 74-kDa regulatory subunits (beta/A and delta/B") in a ratio of 1:1:1. The purified delta/B" was used as an immunogen in mice, to prepare specific antisera against delta/B". Immunoblot analyses with the antisera detected an immunoreactive 72-kDa protein in the cytosol from various rat tissues including erythrocytes, brain, lung, testis, adrenal gland, heart, spleen, kidney, and liver. The 72-kDa protein was highly abundant in brain and was distributed evenly in cerebral cortex, cerebellum, and brain stem. The 72-kDa protein was also detected in mitochondria and microsome fractions. An immunoreactive 68-kDa protein was detected mainly in nuclear and microsome fractions. The 72-kDa protein from rat brain cytosol copurified with phosphorylated H2B histone phosphatase activity during successive chromatographies on DEAE-Toyopearl, AH-Sepharose, Sephadex G-150, H1 histone-Toyopearl, TSK DEAE-5PW, protamine-Toyopearl, and TSK G3000SW columns. The purified enzyme migrated as a single protein band on nondenaturing PAGE and as three protein bands of 34, 63, and 72 kDa in a ratio of 1:1:1 on SDS-PAGE. The molecular weight of the enzyme was estimated to be 170,000 from the s20,W value of 7.2 +/- 0.3 S and the Stokes radius of 5.5 +/- 0.1 nm. The rat brain enzyme was classified as PP2A, based on the following properties; (1) an IC50 for okadaic acid of 10(-9) M; (2) its preferential dephosphorylation of the a subunit of phosphorylase kinase; (3) its insensitivity to protein inhibitor 2; and (4) its heterotrimeric subunit structure. The Km value and the molecular activity of the enzyme for phosphorylated H2B histone were 72.3 +/- 0.3 microM and 192 +/- 2 mol Pi released/min/mol enzyme, respectively, and were comparable to those of human erythrocyte PP2A (alpha1 beta1 delta1/ CAB"). The 72-kDa subunit in the purified rat brain PP2A was phosphorylated in vitro by cAMP-dependent protein kinase.
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PMID:Tissue and subcellular distributions, and characterization of rat brain protein phosphatase 2A containing a 72-kDa delta/B" subunit. 927 86

Human erythrocyte protein phosphatase 2A, which comprises a 34-kDa catalytic C subunit, a 63-kDa regulatory A subunit and a 74-kDa regulatory B'' (delta) subunit, was phosphorylated at serine residues of B'' in vitro by cAMP-dependent protein kinase (A-kinase). In the presence and absence of 0.5 microM okadaic acid (OA), A-kinase gave maximal incorporation of 1.7 and 1.0 mol of phosphate per mol of B'', respectively. The Km value of A-kinase for CAB'' was 0.17 +/- 0.01 microM in the presence of OA. The major in vitro phosphorylation sites of B'' were identified as Ser-60, -75 and -573 in the presence of OA, and Ser-75 and -573 in the absence of OA. Phosphorylation of B'' did not dissociate B'' from CA, and stimulated the molecular activity of CAB'' toward phosphorylated H1 and H2B histones, 3.8- and 1.4-fold, respectively, but not toward phosphorylase a.
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PMID:Activation of protein phosphatase 2A by cAMP-dependent protein kinase-catalyzed phosphorylation of the 74-kDa B'' (delta) regulatory subunit in vitro and identification of the phosphorylation sites. 968 62

At intermediate stages of male pronucleus formation, sperm-derived chromatin is composed of hybrid nucleoprotein particles formed by sperm H1 (SpH1), dimers of sperm H2A-H2B (SpH2A-SpH2B), and a subset of maternal cleavage stage (CS) histone variants. At this stage in vivo, the CS histone variants are poly(ADP-ribosylated), while SpH2B and SpH1 are phosphorylated. We have postulated previously that the final steps of sperm chromatin remodeling involve a cysteine-protease (SpH-protease) that degrades sperm histones in a specific manner, leaving the maternal CS histone variants unaffected. More recently we have reported that the protection of CS histones from degradation is determined by the poly(ADP-ribose) moiety of these proteins. Because of the selectivity displayed by the SpH-protease, the coexistence of a subset of SpH together with CS histone variants at intermediate stages of male pronucleus remodeling remains intriguing. Consequently, we have investigated the phosphorylation state of SpH1 and SpH2B in relation to the possible protection of these proteins from proteolytic degradation. Histones H1 and H2B were purified from sperm, phosphorylated in vitro using the recombinant alpha-subunit of casein kinase 2, and then used as substrates in the standard assay of the SpH-protease. The phosphorylated forms of SpH1 and SpH2B were found to remain unaltered, while the nonphosphorylated forms were degraded. On the basis of this result, we postulate a novel role for the phosphorylation of SpH1 and SpH2B that occurs in vivo after fertilization, namely to protect these histones against degradation at intermediate stages of male chromatin remodeling.
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PMID:Phosphorylation protects sperm-specific histones H1 and H2B from proteolysis after fertilization. 1061 34

Human cytomegalovirus UL97 is an unusual protein kinase that can phosphorylate nucleoside analogs such as ganciclovir but whose specificity for exogenous protein substrates has remained unknown. We found that purified, recombinant glutathione S-transferase-UL97 fusion protein can phosphorylate histone H2B. Phosphorylation was abrogated by substitution of glutamine for a conserved lysine in subdomain II and inhibited by a new antiviral drug, maribavir. Sequencing and mass spectrometric analyses of purified (32)P-labeled tryptic peptides of H2B revealed that the sites of phosphorylation were, in order of extent, Ser-38, Ser-87, Ser-6, Ser-112, and Ser-124. Phosphorylation of synthetic peptides containing these sites, analyzed using a new, chimeric gel system, correlated with their phosphorylation in H2B. Phosphorylation of the Ser-38 peptide by UL97 occurred on Ser-38 and was specifically sensitive to maribavir, whereas phosphorylation of this peptide by cAMP-dependent protein kinase occurred on Ser-36. The extent of phosphorylation was greatest with peptides containing an Arg or Lys residue 5 positions downstream (P+5) from the Ser. Substitution with Ala at this position essentially eliminated activity. These results identify exogenous protein and peptide substrates of UL97, reveal an unusual dependence on the P+5 position, and may abet discovery of new inhibitors of UL97 and human cytomegalovirus replication.
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PMID:Specific phosphorylation of exogenous protein and peptide substrates by the human cytomegalovirus UL97 protein kinase. Importance of the P+5 position. 1204 83


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