Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.11.11 (AMPK)
 12,425 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Ribonucleotide reductase is responsible for supplying the deoxyribonucleotides required for DNA synthesis and repair. The active enzyme consists of two dissimilar protein components called R1 and R2. Immunoprecipitation of R1 and R2 proteins from [32P]orthophosphate-labeled exponentially growing mouse L cells showed that the R2 protein but not the R1 protein of ribonucleotide reductase could be phosphorylated in vivo. Two-dimensional phosphopeptide mapping experiments of trypsin-digested R2 protein showed a major spot containing more than 90% of the total radioactivity and a minor spot with the remaining radioactivity. Phosphoamino acid analysis of R2 phosphorylated protein indicated that phosphorylation occurred exclusively on serine. Protein kinase C, cAMP-dependent protein kinase, p34cdc2, and CDK2 were capable of phosphorylating the R2 protein in vitro, whereas casein kinase II was not. To determine whether any of these enzymes could phosphorylate peptides observed to be phosphorylated in actively growing cells, tryptic phosphopeptide maps of R2 that had been phosphorylated in vitro were compared with maps of R2 that had been isolated from [32P]-labeled cells. Only the phosphopeptide maps obtained with p34cdc2 and CDK2 matched the pattern found in [32P]-labeled cells. Experiments in which tryptic digests from different samples were mixed prior to two-dimensional separation demonstrated comigration of phosphopeptides obtained by in vivo phosphorylation with phosphopeptides derived from p34cdc2 or CDK2 obtained by in vitro phosphorylations. These studies indicate that protein R2 phosphorylation may play an important role in the regulation of ribonucleotide reduction, DNA synthesis, and cell cycle progression, and suggest a potentially important p34cdc2 and/or CDK2 regulation point in DNA replication.
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PMID:Phosphorylation of ribonucleotide reductase R2 protein: in vivo and in vitro evidence of a role for p34cdc2 and CDK2 protein kinases. 825 5

During meiotic maturation or after fertilization of invertebrate and vertebrate oocytes, many of the quiescent stored mRNAs are recruited into polysomes. In the clam, Spisula solidissima, such masked messages include the abundant mRNAs encoding cyclin A and the small subunit of ribonucleotide reductase. We have previously shown that mRNA-specific unmasking of these two messages can be achieved in vitro, in oocyte cell-free extracts, by the addition of antisense RNAs corresponding to a fairly short (130-140 nucleotides) segment in their cognate 3' untranslated regions. We postulated that the antisense RNAs prevented the binding of a masking repressor protein (Standart et al., 1990). Here we report UV-crosslinking and gel retardation studies which show that the masking portions of the translationally regulated mRNAs bind an oocyte protein of 82 kDa (p82), which is phosphorylated after fertilization. This modification was accompanied by altered RNP complex formation in gel retardation assays. These changes presumably reflect the activation of translation of the masked mRNAs. The role of p82 phosphorylation in maternal mRNA unmasking was assessed in a novel in vitro activation system developed from clam oocytes, based upon the natural rise in pH which accompanies fertilization. Concomitant with mRNA unmasking, several kinases, including cdc2 and MAP kinases were activated in this system, as was p82 phosphorylation. Inhibitors of serine/threonine kinases, including 6-DMAP, staurosporine, and H7 inhibited p82 phosphorylation, whereas inhibitors of tyrosine kinases, protein kinase C, cAMP-dependent protein kinase, and p70s6k did not prevent this modification. A specific inhibitor of cdc2 kinase, p27Kip1, prevented p82 phosphorylation and translational activation, strongly suggesting that p82 modification is required for unmasking.
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PMID:Unmasking mRNA in clam oocytes: role of phosphorylation of a 3' UTR masking element-binding protein at fertilization. 857 30

Three ATP-dependent enzymes with different folds, cAMP-dependent protein kinase, D-Ala:D-Ala ligase and the alpha-subunit of the alpha2beta2 ribonucleotide reductase, have a similar organization of their ATP-binding sites. The most meaningful similarity was found over 23 structurally equivalent residues in each protein and includes three strands each from their beta-sheets, in addition to a connecting loop. The equivalent secondary structure elements in each of these enzymes donate four amino acids forming key hydrogen bonds responsible for the common orientation of the "AMP" moieties of their ATP-ligands. One lysine residue conserved throughout the three families binds the alpha-phosphate in each protein. The common fragments of structure also position some, but not all, of the equivalent residues involved in hydrophobic contacts with the adenine ring. These examples of convergent evolution reinforce the view that different proteins can fold in different ways to produce similar structures locally, and nature can take advantage of these features when structure and function demand it, as shown here for the common mode of ATP-binding by three unrelated proteins.
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PMID:Enzyme-mononucleotide interactions: three different folds share common structural elements for ATP recognition. 1008 73

ATP is a ligand common to many proteins, yet it is unclear whether common recognition patterns do exist among the many different folds that bind ATP. Previously, it was shown that cAMP-dependent protein kinase, D-Ala:D-Ala ligase and the alpha-subunit of the alpha 2 beta 2 ribonucleotide reductase do share extensive common structural elements for ATP recognition although their folds are different. Here, we have made a survey of structures that bind ATP and compared them with the key features seen in these three proteins. Our survey shows that 12 different fold types share a specific recognition pattern for the adenine moiety, and 8 of these folds have a common structural framework for recognition of the AMP moiety of the ligand. The common framework consists of a tripeptide segment plus three additional residues, which provides similar polar and hydrophobic interactions between the protein and mononucleotide. Consensus interactions are represented by four key hydrogen bonds present in each fold type. Two of these four hydrogen bonds, together with three aliphatic residues, form a specific recognition pattern for the adenine moiety in all 12 folds. These similarities point to a structural-functional requirement shared by these different mononucleotide-binding proteins that represent at this time 28% of the adenine mononucleotide complexes found in the Brookhaven Protein Data Bank.
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PMID:When fold is not important: a common structural framework for adenine and AMP binding in 12 unrelated protein families. 1071 91

We report the results of phosphoproteomic analysis of mouse thymoma cells treated with tributyltin oxide (TBTO), an immunotoxic compound. After cell lysis, phosphoproteins were isolated using Phosphoprotein Purification Kit, separated by SDS-PAGE and subsequently digested with trypsin. Phosphopeptides were enriched employing titanium dioxide, and the obtained fractions were analyzed by nano-LC-MS/MS. A total of 160 phosphoproteins and 328 phosphorylation sites were identified in thymoma cells. Among the differentially phosphorylated proteins identified in TBTO-treated cells were key enzymes, which catalyze rate-limiting steps in pathways that are sensitive to cellular energy status. These proteins included acetyl-CoA carboxylase isoform 1, which catalyzes the rate-limiting step of fatty acid synthesis. Another enzyme was glutamine: fructose-6-phosphate amidotransferase, GFAT1, the first and rate-limiting enzyme for the hexoamine synthesis pathway. Pyruvate dehydrogenase (PDH), a multicomplex enzyme that catalyzes the rate-limiting step of aerobic oxidation of fuel carbohydrates, was identified in both TBTO-treated and control cells; however, phosphorylation at residue S293, known to inhibit PDH activity, was identified only in control cells. A lower expression level of ribosomal protein S6 kinase 1, a downstream kinase of the mammalian target of rapamycin signaling pathway implicated in protein synthesis through phosphorylation of 40 ribosomal S6, was observed in the treated cells. Giant kinases like AMP-activated protein kinase (AMPK) and cAMP-dependent protein kinase (PKAR1A), which are known to mediate the phosphorylation of these enzymes, were identified in TBTO-treated cells. Downregulation of proteins, such as MAPK, matrin-3 and ribonucleotide reductase, subunit RRM2, which are implicated in cell proliferation, was also observed in TBTO-treated cells. Together, the results show that TBTO affects proliferation and energy sensor pathways.
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PMID:Phosphoproteomic analysis of mouse thymoma cells treated with tributyltin oxide: TBTO affects proliferation and energy sensing pathways. 2217 45