-
The Journal of Biological Chemistry Apr 1987DNA precursor imbalances are known to be mutagenic in both eukaryotic and prokaryotic systems. Almost certainly, such mutagenesis involves competition between correctly...
DNA precursor imbalances are known to be mutagenic in both eukaryotic and prokaryotic systems. Almost certainly, such mutagenesis involves competition between correctly and incorrectly base-paired precursors at replication sites. Since other factors may be involved, it is important to identify specific mutations induced by specific pool imbalances. Using bacteriophage T4, we have developed a system for such analysis. We prepare double mutants of T4; one mutation affects a phage-coded enzyme of deoxyribonucleoside triphosphate (dNTP) metabolism, while the second is an rII mutation known to revert along a specific pathway. We determine dNTP pools in infection by such a mutant and measure both the spontaneous reversion rate of the rII mutation and, in some cases, the nucleotide sequence at the mutant site. In this paper we analyze mutations induced by a deficiency of T4-encoded deoxycytidylate deaminase. This causes pools of 5-hydroxymethyl-dCTP to expand some 30-fold, while dTTP pools contract. This specifically stimulates AT-to-GC reversion. One of the four AT-to-GC reverters tested, rIIUV215, increases its reversion rate at least 1000-fold under these pool-imbalance conditions, while the other mutants tested show increases of only about 10-fold. Therefore, factors other than dNTP competition, including local DNA sequence environment, must be invoked to fully explain mechanisms of dNTP pool imbalance-induced mutagenesis. We discuss models for this, and we also report unexpected effects of the dCMP deaminase deficiency upon pools of ribonucleoside triphosphates.
Topics: Base Sequence; DCMP Deaminase; Deoxyribonucleotides; Escherichia coli; Genes; Genes, Viral; Mutation; Nucleotide Deaminases; T-Phages
PubMed: 3553179
DOI: No ID Found -
Progress in Nucleic Acid Research and... 1990
Review
Topics: Amino Acid Sequence; Base Sequence; DCMP Deaminase; Molecular Sequence Data; Thymidylate Synthase
PubMed: 2247612
DOI: 10.1016/s0079-6603(08)60623-6 -
Journal of Molecular Biology Mar 20082'-Deoxycytidylate deaminase [or deoxycytidine-5'-monophosphate (dCMP) deaminase, dCD] catalyzes the deamination of dCMP to deoxyuridine-5'-monophosphate to provide the...
2'-Deoxycytidylate deaminase [or deoxycytidine-5'-monophosphate (dCMP) deaminase, dCD] catalyzes the deamination of dCMP to deoxyuridine-5'-monophosphate to provide the main nucleotide substrate for thymidylate synthase, which is important in DNA synthesis. The activity of this homohexameric enzyme is allosterically regulated by deoxycytidine-5'-triphosphate (dCTP) as an activator and by deoxythymidine-5'-triphosphate as an inhibitor. In this article, we report the crystal structures of dCMP deaminase from Streptococcus mutans and its complex with dCTP and an intermediate analog at resolutions of 3.0 and 1.66 A. The protein forms a hexamer composed of subunits adopting a three-layer alpha/beta/alpha sandwich fold. The positive allosteric regulator dCTP mainly binds at the interface between two monomers in a molar ratio of 1:1 and rearranges the neighboring interaction networks. Structural comparisons and sequence alignments revealed that dCMP deaminase from Streptococcus mutans belongs to the cytidine deaminase superfamily, wherein the proteins exhibit a similar catalytic mechanism. In addition to the two conserved motifs involved in the binding of Zn(2+), a new conserved motif, (G(43)YNG(46)), related to the binding of dCTP was also identified. N-terminal Arg4, a key residue located between two monomers, binds strongly to the gamma phosphate group of dCTP. The regulation signal was transmitted by Arg4 from the allosteric site to the active site via modifications in the interactions at the interface where the substrate-binding pocket was involved and the relocations of Arg26, His65, Tyr120, and Arg121 to envelope the active site in order to stabilize substrate binding in the complex. Based on the enzyme-regulator complex structure observed in this study, we propose an allosteric mechanism for dCD regulation.
Topics: Allosteric Regulation; Amino Acid Sequence; Binding Sites; Catalysis; Crystallography, X-Ray; DCMP Deaminase; Deoxycytosine Nucleotides; Dimerization; Kinetics; Magnesium; Models, Molecular; Molecular Sequence Data; Protein Structure, Quaternary; Sequence Alignment; Sequence Homology, Amino Acid; Software; Streptococcus mutans; Substrate Specificity
PubMed: 18255096
DOI: 10.1016/j.jmb.2007.12.064 -
Fundamental & Clinical Pharmacology Apr 2011Deoxycytidine analogs (dCa's) are nucleosides widely used in anticancer and anti (retro) viral therapies. Intracellularly phosphorylated dCa anabolites are considered to... (Review)
Review
Deoxycytidine analogs (dCa's) are nucleosides widely used in anticancer and anti (retro) viral therapies. Intracellularly phosphorylated dCa anabolites are considered to be their main active metabolites. This article reviews the literature on the formation and pharmacological activity of deaminated dCa nucleotides. Most dCa's are rapidly deaminated into deoxyuridine analogs (dUa's) which are only slowly phosphorylated and therefore relatively inactive. dUa nucleotides are, however, also formed via deamination of dCa monophosphates by deoxycytidine monophosphate deaminase (dCMPD). dUa-monophosphates can interact with thymidylate synthase (TS), whereas dUa-triphosphates are incorporated into nucleic acids and interfere with polymerases. Administration of dCa's as monophosphate prodrugs or co-administration of the cytidine deaminase inhibitor tetrahydrouridine (THU) does not prevent dUa nucleotide formation which is, on the other hand, influenced by the dose and dCMPD activity. Taken together, these observations show that the formation of dUa nucleotides is a common phenomenon in treatment with dCa's and these compounds may play a role in treatment outcome. We conclude that more attention should be given to these relatively unknown, but potentially important metabolites.
Topics: Animals; Antineoplastic Agents; Antiviral Agents; DCMP Deaminase; Deamination; Deoxycytidine; Deoxyuracil Nucleotides; Dose-Response Relationship, Drug; Humans; Phosphorylation
PubMed: 20199587
DOI: 10.1111/j.1472-8206.2010.00823.x -
Journal of Molecular Biology Sep 1978
Topics: Aldehydes; Binding Sites; DCMP Deaminase; Deoxyadenine Nucleotides; Deoxycytosine Nucleotides; Electrophoresis, Polyacrylamide Gel; Enzyme Activation; Glutaral; Kinetics; Nucleotide Deaminases; Protein Conformation
PubMed: 101672
DOI: 10.1016/0022-2836(78)90152-3 -
Trends in Genetics : TIG Oct 1996RNA editing is a co- or post-transcriptional process in which select nucleotide sequences in RNA are altered from that originally encoded in the genome. The mRNAs... (Review)
Review
RNA editing is a co- or post-transcriptional process in which select nucleotide sequences in RNA are altered from that originally encoded in the genome. The mRNAs encoding apolipoprotein B and some glutamate receptor subunits of ionotropic membrane channels are edited by site-specific base-deamination systems. Although these editing systems differ markedly in their mechanism for RNA-substrate binding and in their catalytic subunits, recent results suggest potentially common solutions to the problem of editing-site selectivity. The data suggest that there are multiple editing complexes or 'editosomes', which manifest editing-site preferences due to their macromolecular composition.
Topics: Amino Acid Sequence; Animals; Apolipoproteins B; Base Sequence; Consensus Sequence; Cytidine Deaminase; DCMP Deaminase; Escherichia coli; Gene Expression Regulation; Glucuronidase; Humans; Mammals; RNA Editing; RNA, Messenger
PubMed: 8909139
DOI: 10.1016/0168-9525(96)10042-1 -
Biochemical Genetics Apr 1975Disc polyacrylamide gel electrophoresis (disc PAGE) analyses have revealed that mouse, human, and monkey cytosal deoxycytidylate (dCMP) deaminases differ in...
Disc polyacrylamide gel electrophoresis (disc PAGE) analyses have revealed that mouse, human, and monkey cytosal deoxycytidylate (dCMP) deaminases differ in electrophoretic mobility, so that mixtures of mouse and human, mouse and monkey and human and monkey enzymes can be separated. To learn whether the genes for dCMP deaminase and thymidine (dT) kinase are genetically linked, disc PAGE analyses of cytosol fractions from human-mouse and monkey-mouse somatic cell hybrids were carried out. The interspecific somatic cell hybrids were derived from the fusion of cytosol dT kinase deficient mouse cells with cytosol dT kinase-positive human and monkey cells: they contained mostly mouse chromosomes and a few primate chromosomes, including the determinant for primate cytosol dT kinase. The disc PAGE analyses demonstrated that the human-mouse and monkey-mouse somatic cell hybrids contained a dCMP deaminase activity with an electrophoretic mobility characteristic of mouse dCMP deaminase. Enzymes with electrophoretic mobilities characteristic of human and monkey dCMP deaminases were not demonstrable. these findings suggest that primate cytosol dT kinase and dCMP deaminase are coded on different chromosomes, or that the formation in hybird cells of an active primate dCMP deaminase is suppressed. Chick-mouse somatic cell hybrids containing chick but not mouse cytosol dT kinase were also analyzed, but it was not possible to establish whether the enzyme was of murine or avian origin because of the similarity in electrophoretic mobility between the chick and mouse enzymes. Human and mouse cells contained low levels of mitochondrial dCMP deaminase activity. In contrast to dT kinase isozymes, however, mitochondrial and cytosol dCMP deaminases were electrophoretically indistinguishable.
Topics: Aminohydrolases; Animals; Cell Fusion; Centrifugation, Density Gradient; Chromosome Mapping; Clone Cells; Electrophoresis, Polyacrylamide Gel; Genes; Genetic Linkage; Haplorhini; Humans; Hybrid Cells; In Vitro Techniques; Isoenzymes; Mice; Mitochondria; Species Specificity; Thymidine Kinase
PubMed: 807195
DOI: 10.1007/BF00486015 -
British Journal of Obstetrics and... Apr 1982A simple and reliable method for the estimation of cytidine deaminase (cytidine aminohydrolase; EC 3.5.4.5) activity in pregnancy serum is described. This enzyme does...
A simple and reliable method for the estimation of cytidine deaminase (cytidine aminohydrolase; EC 3.5.4.5) activity in pregnancy serum is described. This enzyme does not require magnesium for activation and is also more stable than deoxycytidylate deaminase (dCMP deaminase, dCMP aminohydrolase; EC 3.5.4.12). There was excellent correlation between the two enzymes (r = 0.92). Both enzymes showed increased activity in abnormal pregnancy. Both enzymes activities were found to be similar in 1305 maternal serum samples and therefore due to the simplicity of cytidine deaminase estimation it is recommended for the screening of large numbers of antenatal sera for abnormal pregnancies.
Topics: Cytidine Deaminase; DCMP Deaminase; Enzyme Activation; Female; Humans; Magnesium; Male; Nucleoside Deaminases; Pre-Eclampsia; Pregnancy; Pregnancy Trimester, Third
PubMed: 7073999
DOI: 10.1111/j.1471-0528.1982.tb04703.x -
British Medical Journal Apr 1975Deoxycytidylate (DCMP) deaminase was assayed at various times during and after normal and abnormal pregnancies. The level in amniotic fluid was assessed at induction and... (Comparative Study)
Comparative Study
Deoxycytidylate (DCMP) deaminase was assayed at various times during and after normal and abnormal pregnancies. The level in amniotic fluid was assessed at induction and at caesarean section, and cord blood levels were estimated after normal delivery and at caesarean section. A rise occurred during labour and after hysterectomy and caesarean section--returning to normal after 2-3, and 12 days respectively. Levels above 4.8 X 10-minus 4 ml-minus 1 were found in cases of preeclamptic toxaemia and early intrauterine death and in twin pregnancies over 36 weeks' gestation. It is suggested that because of its low incidence of false-negative and false-positive results this test is far superior to other enzyme tests in pregnancy, and a further trial is in progress to assess its role.
Topics: Aminohydrolases; Amniotic Fluid; Blood; Cesarean Section; Clinical Enzyme Tests; Deoxycytidine Monophosphate; Female; Fetal Death; Humans; Hysterectomy; Labor, Induced; Labor, Obstetric; Male; Pre-Eclampsia; Pregnancy; Pregnancy Complications; Pregnancy, Multiple; Umbilical Cord
PubMed: 1137771
DOI: 10.1136/bmj.2.5961.10 -
Annals of Neurology May 2017Thymidine kinase 2 (TK2), a critical enzyme in the mitochondrial pyrimidine salvage pathway, is essential for mitochondrial DNA (mtDNA) maintenance. Mutations in the...
OBJECTIVE
Thymidine kinase 2 (TK2), a critical enzyme in the mitochondrial pyrimidine salvage pathway, is essential for mitochondrial DNA (mtDNA) maintenance. Mutations in the nuclear gene, TK2, cause TK2 deficiency, which manifests predominantly in children as myopathy with mtDNA depletion. Molecular bypass therapy with the TK2 products, deoxycytidine monophosphate (dCMP) and deoxythymidine monophosphate (dTMP), prolongs the life span of Tk2-deficient (Tk2 ) mice by 2- to 3-fold. Because we observed rapid catabolism of the deoxynucleoside monophosphates to deoxythymidine (dT) and deoxycytidine (dC), we hypothesized that: (1) deoxynucleosides might be the major active agents and (2) inhibition of deoxycytidine deamination might enhance dTMP+dCMP therapy.
METHODS
To test these hypotheses, we assessed two therapies in Tk2 mice: (1) dT+dC and (2) coadministration of the deaminase inhibitor, tetrahydrouridine (THU), with dTMP+dCMP.
RESULTS
We observed that dC+dT delayed disease onset, prolonged life span of Tk2-deficient mice and restored mtDNA copy number as well as respiratory chain enzyme activities and levels. In contrast, dCMP+dTMP+THU therapy decreased life span of Tk2 animals compared to dCMP+dTMP.
INTERPRETATION
Our studies demonstrate that deoxynucleoside substrate enhancement is a novel therapy, which may ameliorate TK2 deficiency in patients. Ann Neurol 2017;81:641-652.
Topics: Animals; Antimetabolites; DNA, Mitochondrial; Deoxycytidine Monophosphate; Disease Models, Animal; Drug Therapy, Combination; Metabolism, Inborn Errors; Mice; Mice, Transgenic; Mitochondrial Diseases; Tetrahydrouridine; Thymidine; Thymidine Kinase
PubMed: 28318037
DOI: 10.1002/ana.24922