Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.7.7 (DNA polymerase)
 17,007 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The structure of Mycobacterium tuberculosis dUTP nucleotidohydrolase (dUTPase) has been determined at 1.3 Angstrom resolution in complex with magnesium ion and the non-hydrolyzable substrate analog, alpha,beta-imido dUTP. dUTPase is an enzyme essential for depleting potentially toxic concentrations of dUTP in the cell. Given the importance of its biological role, it has been proposed that inhibiting M.tuberculosis dUTPase might be an effective means to treat tuberculosis infection in humans. The crystal structure presented here offers some insight into the potential for designing a specific inhibitor of the M.tuberculosis dUTPase enzyme. The structure also offers new insights into the mechanism of dUTP hydrolysis by providing an accurate representation of the enzyme-substrate complex in which both the metal ion and dUTP analog are included. The structure suggests that inclusion of a magnesium ion is important for stabilizing the position of the alpha-phosphorus for an in-line nucleophilic attack. In the absence of magnesium, the alpha-phosphate of dUTP can have either of the two positions which differ by 4.5 Angstrom. A transiently ordered C-terminal loop further assists catalysis by shielding the general base, Asp83, from solvent thus elevating its pK(a) so that it might in turn activate a tightly bound water molecule for nucleophilic attack. The metal ion coordinates alpha, beta, and gamma phosphate groups with tridentate geometry identical with that observed in the crystal structure of DNA polymerase beta complexed with magnesium and dNTP analog, revealing some common features in catalytic mechanism.
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PMID:Crystal structure of the Mycobacterium tuberculosis dUTPase: insights into the catalytic mechanism. 1527 40

Ostreid herpesvirus 1 (OsHV-1) is the only member of the Herpesviridae that has an invertebrate host and is associated with sporadic mortality in the Pacific oyster (Crassostrea gigas) and other bivalve species. Cryo-electron microscopy of purified capsids revealed the distinctive T=16 icosahedral structure characteristic of herpesviruses, although the preparations examined lacked pentons. The gross genome organization of OsHV-1 was similar to that of certain mammalian herpesviruses (including herpes simplex virus and human cytomegalovirus), consisting of two invertible unique regions (U(L), 167.8 kbp; U(S), 3.4 kbp) each flanked by inverted repeats (TR(L)/IR(L), 7.6 kbp; TR(S)/IR(S), 9.8 kbp), with an additional unique sequence (X, 1.5 kbp) between IR(L) and IR(S). Of the 124 unique genes predicted from the 207 439 bp genome sequence, 38 were members of 12 families of related genes and encoded products related to helicases, inhibitors of apoptosis, deoxyuridine triphosphatase and RING-finger proteins, in addition to membrane-associated proteins. Eight genes in three of the families appeared to be fragmented. Other genes that did not belong to the families were predicted to encode DNA polymerase, the two subunits of ribonucleotide reductase, a helicase, a primase, the ATPase subunit of terminase, a RecB-like protein, additional RING-like proteins, an ion channel and several other membrane-associated proteins. Sequence comparisons showed that OsHV-1 is at best tenuously related to the two classes of vertebrate herpesviruses (those associated with mammals, birds and reptiles, and those associated with bony fish and amphibians). OsHV-1 thus represents a third major class of the herpesviruses.
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PMID:A novel class of herpesvirus with bivalve hosts. 1560 30

We determined the sequence of the 152,372 bp genome of phiYS40, a lytic tailed bacteriophage of Thermus thermophilus. The genome contains 170 putative open reading frames and three tRNA genes. Functions for 25% of phiYS40 gene products were predicted on the basis of similarity to proteins of known function from diverse phages and bacteria. phiYS40 encodes a cluster of proteins involved in nucleotide salvage, such as flavin-dependent thymidylate synthase, thymidylate kinase, ribonucleotide reductase, and deoxycytidylate deaminase, and in DNA replication, such as DNA primase, helicase, type A DNA polymerase, and predicted terminal protein involved in initiation of DNA synthesis. The structural genes of phiYS40, most of which have no similarity to sequences in public databases, were identified by mass spectrometric analysis of purified virions. Various phiYS40 proteins have different phylogenetic neighbors, including myovirus, podovirus, and siphovirus gene products, bacterial genes and, in one case, a dUTPase from a eukaryotic virus. phiYS40 has apparently arisen through multiple acts of recombination between different phage genomes as well as through acquisition of bacterial genes.
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PMID:Thermus thermophilus bacteriophage phiYS40 genome and proteomic characterization of virions. 1702 29

Genomic analysis of the hyperthermophilic archaeon Thermococcus onnurineus NA1 (TNA1) revealed the presence of a 471-bp open reading frame with 93% similarity to the dUTPase from Pyrococcus furiosus. The dUTPase-encoding gene was cloned and expressed in Escherichia coli. The purified protein hydrolyzed dUTP at about a 10-fold higher rate than dCTP. The protein behaved as a dimer in gel filtration chromatography, even though it contains five motifs that are conserved in all homotrimeric dUTPases. The dUTPase showed optimum activity at 80 degrees C and pH 8.0, and it was highly thermostable with a half-life (t (1/2)) of 170 min at 95 degrees C. The enzymatic activity of the dUTPase was largely unaffected by variations in MgCl(2), KCl, (NH(4))(2)SO(4), and Triton X-100 concentrations, although it was reduced by bovine serum albumin. Addition of the dUTPase to polymerase chain reactions (PCRs) run with TNA1 DNA polymerase significantly increased product yield, overcoming the inhibitory effect of dUTP. Further, addition of the dUTPase allowed PCR amplification of targets up to 15 kb in length using TNA1 DNA polymerase. This enzyme also improved the PCR efficiency of other archaeal family B type DNA polymerases, including Pfu and KOD.
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PMID:Characterization of a dUTPase from the hyperthermophilic archaeon Thermococcus onnurineus NA1 and its application in polymerase chain reaction amplification. 1754 47

Psychological stress-associated immune dysregulation has been shown to disrupt the steady-state expression and reactivate latent herpes viruses. One such virus is the Epstein Barr virus (EBV), which is associated with several human malignancies. EBV infects >90% of people living in North America and persists for life in latently infected cells. Although several studies have shown that glucocorticoids (GCs) can directly induce reactivation of the latent virus, the mechanism of stress hormone involvement in the control of EBV gene expression is not well understood. In this study, we tested the hypothesis that GCs can induce the latent EBV genome to lytically replicate through the induction of the EBV immediate early gene BZLF1 which encodes the lytic transactivator protein ZEBRA. We show a dose-dependent upregulation of BZLF1 mRNA expression by hydrocortisone (HC) and dexamethasone (Dex) in Daudi cells, an EBV genome positive Burkitt's lymphoma cell line, and Dex-induction of the early gene products BLLF3 (encoding for the EBV dUTPase) and BALF5 (encoding for the EBV DNA polymerase). We show that Daudi cells express glucocorticoid receptors (GR) that mediate Dex-dependent upregulation of BZLF1 mRNA levels. This effect was inhibited by both the glucocorticoid receptor antagonist RU486 and by cycloheximide. The results suggest that GCs, in addition to inducing stress-related immune dysregulation, can mediate latent EBV reactivation through the induction of the BZLF1 gene.
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PMID:Glucocorticoids activate Epstein Barr virus lytic replication through the upregulation of immediate early BZLF1 gene expression. 2046 55


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