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Proceedings of the National Academy of... May 1964
Topics: Adenine Nucleotides; Adenosine Triphosphate; Azotobacter; Chromatography; Cytosine Nucleotides; DNA; DNA, Bacterial; Deoxyribonucleases; Guanine Nucleotides; Ligases; Metabolism; Poly A; RNA; RNA, Bacterial; Research; Transferases
PubMed: 14173002
DOI: 10.1073/pnas.51.5.853 -
Structure-Activity Relationship of 3-Methylcytidine-5'-α,β-methylenediphosphates as CD73 Inhibitors.Journal of Medicinal Chemistry Feb 2022We recently reported -substituted 3-methylcytidine-5'-α,β-methylenediphosphates as CD73 inhibitors, potentially useful in cancer immunotherapy. We now expand the...
We recently reported -substituted 3-methylcytidine-5'-α,β-methylenediphosphates as CD73 inhibitors, potentially useful in cancer immunotherapy. We now expand the structure-activity relationship of pyrimidine nucleotides as human CD73 inhibitors. 4-Chloro (MRS4598 ; = 0.673 nM) and 4-iodo (MRS4620 ; = 0.436 nM) substitution of the -benzyloxy group decreased by ∼20-fold. Primary alkylamine derivatives coupled through a -amido group with a varying methylene chain length ( and ) were functionalized congeners, for subsequent conjugation to carrier or reporter moieties. X-ray structures of hCD73 with two inhibitors indicated a ribose ring conformational adaptation, and the benzyloxyimino group ( configuration) binds to the same region (between the C-terminal and N-terminal domains) as -benzyl groups in adenine inhibitors. Molecular dynamics identified stabilizing interactions and predicted conformational diversity. Thus, by -benzyloxy substitution, we have greatly enhanced the inhibitory potency and added functionality enabling molecular probes. Their potential as anticancer drugs was confirmed by blocking CD73 activity in tumor tissues in situ.
Topics: 5'-Nucleotidase; Adult; Cytosine Nucleotides; Diphosphonates; Enzyme Inhibitors; GPI-Linked Proteins; Humans; Male; Molecular Docking Simulation; Molecular Dynamics Simulation; Molecular Structure; Neoplasms; Palatine Tonsil; Protein Binding; Structure-Activity Relationship
PubMed: 35080883
DOI: 10.1021/acs.jmedchem.1c01852 -
Nucleic Acids Research Jan 1977Viroids are uncoated infectious RNA molecules (MW 107 000-127 000) known as pathogens of certain higher plants. Thermodynamic and kinetic studies were carried out on...
Viroids are uncoated infectious RNA molecules (MW 107 000-127 000) known as pathogens of certain higher plants. Thermodynamic and kinetic studies were carried out on highly purified viroid preparations by applying UV-absorption melting analysis and temperature jump methods. The thermal denaturation of viroids is characterized by high thermal stability, high cooperativity and a high degree of base pairing. Two relaxation processes could be resolved; a process in the sec range could be evaluated as an independent all-or-none-transition with the following properties: reaction enthalpy= 550 kcal/mol, activation enthalpy of the dissociation = 470 kcal/mol; G : C content = 72 %. These data indicate the existence of an uninterrupted double helix of 52 base pairs. A process in the msec range involves 15 - 25 base pairs which are most probably distributed over several short double helical stretches. A tentative model for the secondary structure of viroids isproposed and the possible functional implications of their physicochemical properties are discussed.
Topics: Chemical Phenomena; Chemistry; Cytosine Nucleotides; Guanine Nucleotides; Kinetics; Nucleic Acid Conformation; Nucleic Acid Denaturation; Plant Diseases; Plants; RNA; Ribonucleotides; Temperature; Thermodynamics; Urea
PubMed: 866174
DOI: 10.1093/nar/4.1.177 -
Molecular Plant Jul 2013Transcription activator-like effectors (TALEs) from Xanthomonas sp. have been used as customizable DNA-binding modules for genome-engineering applications. Ralstonia...
Transcription activator-like effectors (TALEs) from Xanthomonas sp. have been used as customizable DNA-binding modules for genome-engineering applications. Ralstonia solanacearum TALE-like proteins (RTLs) exhibit similar structural features to TALEs, including a central DNA-binding domain composed of 35 amino acid-long repeats. Here, we characterize the RTLs and show that they localize in the plant cell nucleus, mediate DNA binding, and might function as transcriptional activators. RTLs have a unique DNA-binding architecture and are enriched in repeat variable di-residues (RVDs), which determine repeat DNA-binding specificities. We determined the DNA-binding specificities for the RVD sequences ND, HN, NP, and NT. The RVD ND mediates highly specific interactions with C nucleotide, HN interacts specifically with A and G nucleotides, and NP binds to C, A, and G nucleotides. Moreover, we developed a highly efficient repeat assembly approach for engineering RTL effectors. Taken together, our data demonstrate that RTLs are unique DNA-targeting modules that are excellent alternatives to be tailored to bind to user-selected DNA sequences for targeted genomic and epigenomic modifications. These findings will facilitate research concerning RTL molecular biology and RTL roles in the pathogenicity of Ralstonia spp.
Topics: Amino Acid Sequence; Bacterial Proteins; Base Sequence; Cytosine Nucleotides; DNA; Molecular Sequence Data; Ralstonia; Repetitive Sequences, Amino Acid; Substrate Specificity; Transcriptional Activation
PubMed: 23300258
DOI: 10.1093/mp/sst006 -
Cell Dec 2000
Review
Topics: Animals; Bacterial Infections; Cytosine Nucleotides; DNA Methylation; Guanine Nucleotides; Immune System
PubMed: 11163174
DOI: 10.1016/s0092-8674(00)00201-4 -
The Biochemical Journal Jan 19671. The effects of the administration of dl-ethionine on some aspects of lipid and nucleotide metabolism in rat liver were studied. 2. In ethionine-treated animals...
1. The effects of the administration of dl-ethionine on some aspects of lipid and nucleotide metabolism in rat liver were studied. 2. In ethionine-treated animals neutral fat was increased, whereas phospholipids and cholesterol were unchanged. Lipogenesis in vitro was inhibited. 3. The concentration of nicotinamide nucleotides, purine nucleotides and pyrimidine nucleotides was decreased. The decrease was due to free adenine nucleotides, inosine nucleotides, uridine nucleotides and cytidine nucleotides. Also, the protein-bound biotin content was lower. 4. In biotin-deficient rats the development of ethionine-induced fatty liver was inhibited. 5. The possibility was considered that ethionine might produce an inhibition of the synthesis of biotin-dependent acetyl-CoA carboxylase.
Topics: Acetates; Adenine Nucleotides; Animals; Biotin; Carbon Isotopes; Cholesterol; Cytosine Nucleotides; Ethionine; Fatty Liver; Female; Guanine Nucleotides; In Vitro Techniques; Lipids; Liver; Nucleotides; Phospholipids; Rats; Uracil Nucleotides; Vitamin B Deficiency
PubMed: 6030278
DOI: 10.1042/bj1020163 -
Molecules (Basel, Switzerland) Jul 2021decomposes nicotine through the pyridine pathway. 6-hydroxypseudooxynicotine 2-oxidoreductase (also named ketone dehydrogenase, Kdh) is an important enzyme in nicotine...
decomposes nicotine through the pyridine pathway. 6-hydroxypseudooxynicotine 2-oxidoreductase (also named ketone dehydrogenase, Kdh) is an important enzyme in nicotine degradation pathway of , and is responsible for the second hydroxylation of nicotine. Kdh belongs to the molybdenum hydroxylase family, and catalyzes the oxidation of 6-hydroxy-pseudooxynicotine (6-HPON) to 2,6-dihydroxy-pseudooxynicotine (2,6-DHPON). We determined the crystal structure of the Kdh holoenzyme from , with its three subunits KdhL, KdhM, and KdhS, and their associated cofactors molybdopterin cytosine dinucleotide (MCD), two iron-sulfur clusters (FeS), and flavin adenine dinucleotide (FAD), respectively. In addition, we obtained a structural model of the substrate 6-HPON-bound Kdh through molecular docking, and performed molecular dynamics (MD) and quantum mechanics/molecular mechanics (QM/MM) calculations to unveil the catalytic mechanism of Kdh. The residues Glu345, Try551, and Glu748 of KdhL were found to participate in substrate binding, and Phe269 and Arg383 of KdhL were found to contribute to stabilize the MCD conformation. Furthermore, site-directed mutagenesis and enzymatic activity assays were performed to support our structural and computational results, which also revealed a trend of increasing catalytic efficiency with the increase in the buffer pH. Lastly, our electrochemical results demonstrated electron transfer among the various cofactors of Kdh. Therefore, our work provides a comprehensive structural, mechanistic, and functional study on the molybdenum hydroxylase Kdh in the nicotine degradation pathway of
Topics: Bacterial Proteins; Cloning, Molecular; Cytosine Nucleotides; Micrococcaceae; Mixed Function Oxygenases; Molecular Docking Simulation; Molybdenum; Nicotine; Pterins; Structure-Activity Relationship
PubMed: 34299660
DOI: 10.3390/molecules26144387 -
Journal of Virology Sep 1975To characterize and compare the thymidine (TdR) and deoxycytidine (CdR) kinase isozymes of uninfected and herpesvirus-infected cells: (i) the subcellular distribution of... (Comparative Study)
Comparative Study
To characterize and compare the thymidine (TdR) and deoxycytidine (CdR) kinase isozymes of uninfected and herpesvirus-infected cells: (i) the subcellular distribution of the isozymes has been studied; (ii) a specific assay for CdR kinase has been devised; (iii) the TdR kinase isozymes have been partially purified; and (iv) the purified enzymes have been analyzed by disc polyacrylamide gel electrophoresis, isoelectric focusing, and glycerol gradient centrifugation and by substrate competition and dCTP inhibition studies. The results indicate that there are interesting individual differences with respect to nucleoside acceptor specificity between the cytosol and mitochondrial pyrimidine deoxyribonucleoside kinases of uninfected cells and between the enzymes induced by different herpesviruses. In the cytosol of uninfected mouse, chicken, and owl monkey kidney cells, two different proteins, TdR kinase F and CdR kinase 2, catalyze the phosphorylations of TdR and CdR, respectively. TdR kinase F does not phosphorylate CdR, nor does CdR kinase 2 phosphorylate TdR. A second TdR kinase isozyme present in HeLa(BU25) mitochondria (TdR kinase B) also lacks CdR phosphorylating activity. In contrast, a genetically distinctive deoxypyrimidine kinase (TdR kinase A) of mouse, human, and chick mitochondria catalyzes the phosphorylation of both TdR and CdT. Three herpesviruses, marmoset herpesvirus and herpes simplex virus types 1 and 2, induce in the cytosol fraction of LM(TK-) mouse cells isozymes which share common properties with mitochondrial TdR kinase A, including the ability to catalyze the phosphorylation of both TdR and CdR. However, the herpesvirus-induced deoxypyrimidine kinases differ from mitochondrial TdR kinase A with respect to sedimentation coefficient, sensitivity to dCTP inhibition, and antigenic determinants. The herpesvirus-specific and the mitochondrial deoxypyrimidine kinases exhibit a preference for TdR over CdR as nucleoside acceptor. Pseudorabies virus and herpesvirus of turkeys induce cytosol TdR kinases resembling the other herpesvirus-induced TdR kinases in several properties, but like cellular TdR kinase F, the pseudorabies virus and herpesvirus of turkeys TdR kinases lack detectable CdR phosphorylating activities. Finally, a marmoset herpesvirus nutant resistant to bromodeoxyuridine, equine herpesvirus type 1, and Herpesvirus aotus induces neither TdR nor CdR phosphorylating enzymes during productive infections.
Topics: Cell Line; Cytoplasm; Cytosine Nucleotides; Deoxycytidine; Deoxycytidine Monophosphate; HeLa Cells; Herpesviridae; Herpesvirus 1, Suid; Kinetics; Mitochondria; Phosphotransferases; Simplexvirus; Structure-Activity Relationship; Thymidine Kinase
PubMed: 169387
DOI: 10.1128/JVI.16.3.486-497.1975 -
FEBS Letters Oct 1984Pyrimidine metabolism in Trichomonas vaginalis was investigated using washed cell suspensions of the organism with radiolabelled pyrimidine ring precursors and preformed...
Pyrimidine metabolism in Trichomonas vaginalis was investigated using washed cell suspensions of the organism with radiolabelled pyrimidine ring precursors and preformed pyrimidines. The precursors [14C]orotate, [14C]bicarbonate and [14C]aspartate were not incorporated into the pyrimidine bases of trichomonal nucleic acids, indicating that the protozoan is unable to synthesise the pyrimidine ring and is dependent on the salvage of exogenous pyrimidines. [3H]uracil, [3H]uridine, [3H]cytidine, deoxy[3H]cytidine and [3H]thymidine were all efficiently salvaged, and interconversion between cytosine and uracil nucleotides was detected. Thymidylate synthase activity was not detected, suggesting that T. vaginalis is dependent upon an exogenous supply of thymidine for TMP synthesis.
Topics: Animals; Aspartic Acid; Bicarbonates; Cytosine Nucleotides; Orotic Acid; Pyrimidines; Thymidine; Thymidylate Synthase; Trichomonas vaginalis; Uracil Nucleotides
PubMed: 6333357
DOI: 10.1016/0014-5793(84)80910-2 -
The Journal of Biological Chemistry Mar 1975Poly(A) polymerase activity was first detected in yeast extracts, primarily in association with the ribosomal fraction, by Twu and Bretthauer in 1971 (Twu, J. S., and...
Poly(A) polymerase activity was first detected in yeast extracts, primarily in association with the ribosomal fraction, by Twu and Bretthauer in 1971 (Twu, J. S., and Bretthauer, RK. (1971) Biochemistry 10, 1576-1582). This activity has now been separated into three distinct enzymes by chromatography on DEAE-cellulose. Each of the three enzymes can catalyze the incorporation of adenylate residues from ATP into a polyadenylate (poly(A)) tract at the 3' terminus of a primer RNA. Enzyme I elutes at 0.07 M ammonium sulfate from the DEAE-cellulose column, utilizes the mixed polynucleotide poly(A,G,C,U) or ribosomal RNA most efficiently in vitro, and may be responsible in vivo for the initiation of the poly(A) tracts found on yeast messenger RNA. Enzyme II elutes from the column at 0.20 M ammonium sulfate, requires poly(A) itself or an RNA primer containing a 3'-oligo(A) tract, and may be responsible in the nucleus for the elongation of tracts initiated by enzyme I. Enzyme III elutes from the column at 0.56 M ammonium sulfate and is present in low amounts in nuclear extracts. It may be involved in adding poly(A) tracts to messenger RNA in mitochondria. These enzymes also have the intrinsic capacity for the incorporation of cytidylate residues from CTP, which correlates with the finding of cytidylate residues in the poly(A) tracts present in the yeast RNA which is rapidly labeled in vivo. About 75% of the total poly(A) polymerase activity of yeast is enzyme I, most of which is present in the soluble protein fraction of the whole yeast extract. About 20% of the total poly(A) polymerase is enzyme II, and 1 to 5% is enzyme III. All three of the yeast poly(A) polymerases require an RNA primer with a free 3'-hydroxyl group, show no requirement for a DNA template, require Mn-2+ for optimal activity, have pH optima of 8.5, and are inhibited by GTP, CTP, UTP, and native yeast DNA. Polymerases I and II have similar molecular weights by gel filtration.
Topics: Adenosine Triphosphate; Cytosine Nucleotides; Guanosine Triphosphate; Hydrogen-Ion Concentration; Kinetics; Manganese; Nucleotidyltransferases; Osmolar Concentration; Polynucleotides; RNA, Messenger; Ribosomes; Saccharomyces cerevisiae; Uracil Nucleotides
PubMed: 234467
DOI: No ID Found