Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: 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)

A screening program for antiviral drugs begun at Burroughs Wellcome in the 1960s resulted in the discovery of acyclovir in 1974. Preclinical investigation brought the drug to clinical trials in 1977 and the first form of the drug (topical) was available to physicians in 1982. Activity of acyclovir is greatest against herpes 1 and herpes 2, less against varicella zoster, still less against Epstein-Barr, and very little against cytomegalovirus. Acyclovir is an antiviral agent only after it is phosphorylated in infected cells by a viral-induced thymidine kinase. Acyclovir monophosphate is phosphorylated to diphosphate and triphosphate forms by cellular enzymes in the infected host cell where the drug is concentrated. Acyclovir triphosphate inactivates viral deoxyribonucleic acid polymerase. Acyclovir incorporation into the growing viral deoxyribonucleic acid chain causes its termination. The antiviral process has relatively little effect on normal, uninfected cells. An important toxic effect of acyclovir is its potential to cause obstructive nephropathy. The drug is excreted primarily by the kidney, which may require smaller doses in patients with decreased kidney function. Oral dosages of acyclovir as recommended for herpes simplex are probably not adequate for varicella zoster infections.
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PMID:History, pharmacokinetics, and pharmacology of acyclovir. 282 40

The genome of herpes simplex virus codes for several enzymes, including viral thymidine kinase and viral deoxyribonucleic acid (DNA) polymerase. When viral resistance develops, it does so by changes in these two enzymes. Three possible mechanisms of viral resistance to acyclovir include (1) selection of viral mutants that make little or no thymidine kinase and do not phosphorylate acyclovir adequately, (2) selection of mutants that can phosphorylate thymidine but cannot phosphorylate acyclovir (i.e., these viruses have thymidine kinases with altered substrate specificity), and (3) selection of viruses that have altered DNA polymerases that replicate viral DNA in the presence of acyclovir triphosphate. Thymidine kinase-deficient virus has been isolated from clinical isolates frequently, but few strains appear to be virulent for animals or humans and only a few seem to have caused clinical disease. Viruses with altered substrate specificity have been reported but viruses with an altered DNA polymerase have not occurred in clinical practice. Antiviral drugs should be used only when necessary to minimize the appearance of resistant strains of virus.
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PMID:Significance of resistance of herpes simplex virus to acyclovir. 282 41

The effect of nucleoside-5-triphosphates analogues on the DNA polymerase of herpes simplex virus (HSV) has been investigated. Evidence is obtained that 3-amino-2,3-dideoxythymidine triphosphate selectively inhibits the DNA synthesis, catalyzed by HSV DNA polymerase. 3-amino-2,3-dideoxythymidine exhibits antiherpetic effect in single cells cultures. It may be phosphorylated by cellular thymidine kinase. The nuclei of Vero cells infected by HSV are an adequate system for antiherpetic compounds screening.
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PMID:[Inhibitory effect of various analogs of nucleoside-5'-triphosphates on DNA synthesis catalyzed by DNA polymerase from herpes simplex virus type I]. 282 34

We have mapped the termini and determined the relative abundance and ribosome density of the major cytoplasmic transcript of the DNA polymerase (pol) gene of herpes simplex virus type 1. Nuclease protection and primer extension analyses located the 5' end of the major pol transcript at two closely spaced sites 51 and 57 nucleotides to the left of a BamHI site at map position 0.413. S1-sensitive sites corresponding to additional minor transcripts were found to map further upstream within a palindromic sequence that contains a viral replication origin. The major 3' end was found to map 90 nucleotides upstream of a KpnI site at map position 0.439. Quantitative S1 nuclease assays revealed that pol transcripts were nearly as abundant as transcripts encoded by the viral thymidine kinase gene. However, relatively few pol transcripts were found on large polysomes at 5.5 h after infection, when pol transcripts were most abundant. This was in marked contrast to the polyribosome distribution of transcripts from the thymidine kinase gene and the major DNA-binding protein gene. These results and sequence features of the pol transcript suggest that pol expression is regulated, in part, at the level of translation.
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PMID:Analysis of the transcript of the herpes simplex virus DNA polymerase gene provides evidence that polymerase expression is inefficient at the level of translation. 283 6

A series of analogues of acyclovir and ganciclovir were prepared in which conformational constraints were imposed by incorporation of a cyclopropane ring or unsaturation into the side chain. In addition, several related base-modified compounds were synthesized. These acyclonucleosides were evaluated for enzymatic phosphorylation and DNA polymerase inhibition in a staggered assay and for inhibitory activity against herpes simplex virus types 1 and 2 in vitro. Certain of the guanine or 8-azaguanine derivatives were good substrates for the viral thymidine kinase and were further converted to triphosphate, but none was a potent inhibitor of the viral DNA polymerase. Nevertheless, one member of this group, (+/-)-9-[[(Z)-2-(hydroxymethyl)cyclopropyl]methyl]guanine (3a), displayed significant antiherpetic activity in vitro, superior to that of the corresponding cis olefin 4a. Another group, typified by (+/-)-9-[[(E)-2-(hydroxymethyl)cyclopropyl]methyl]adenine (17b), possessed modest antiviral activity despite an apparent inability to be enzymatically phosphorylated. The relationship of side-chain conformation and flexibility to biological activity in this series is discussed.
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PMID:Synthesis and antiherpetic activity of (+/-)-9-[[(Z)-2-(hydroxymethyl)cyclopropyl]methyl]guanine and related compounds. 284 25

(R,S)-9-(3-hydroxy-2-phosphonomethoxypropyl)guanine [(R,S)-HPMPG] exhibits broad spectrum antiviral activity with an ED50 of less than 1 microM against herpes simplex virus (HSV) types 1 and 2, varicella zoster virus, human cytomegalovirus (HCMV) and vaccinia in plaque reduction assays. Wild type HSV-2 and its thymidine kinase deficient variant are equally sensitive to (R,S)-HPMPG. (R,S)-HPMPG is 100-fold more potent than acyclovir (ED50 = 0.45 microM vs. 44 microM, respectively) against HCMV in cell culture, and 10-fold more active than acyclovir in extending survival time in mice intraperitoneally infected with 70 LD50 HSV-1. However, (R,S)-HPMPG is toxic when administered repeatedly at 44 mg/kg/day in uninfected adult mice. The diphosphoryl derivative of HPMPG was enzymatically synthesized and is a competitive inhibitor of HSV-1 DNA polymerase relative to dGTP (K1 = 0.03 microM). HPMPG-PP is 70-fold less active at inhibiting HeLa DNA polymerase alpha than HSV-1 DNA polymerase. At concentrations between 0.3 and 1.5 microM (R,S)-HPMPG inhibited HSV-1 DNA replication greater than or equal to 50% in infected cells as measured by nucleic acid hybridization. Consistent with inhibition of viral DNA synthesis, 6 to 30 microM (R,S)-HPMPG reduces late viral polypeptide synthesis in HSV-1 infected cells. These data indicate that (R,S)-HPMPG is a thymidine kinase independent broad spectrum antiviral drug which is capable of inhibiting viral DNA polymerase.
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PMID:Broad-spectrum antiviral activity of the acyclic guanosine phosphonate (R,S)-HPMPG. 285 86

We have undertaken a search for mammalian DNA-binding proteins that enhance the activity of DNA polymerases in a template sequence-specific fashion. In this paper, we report the extensive purification and characterization of a new DNA-binding protein from rabbit liver that selectively stimulates DNA polymerases to copy synthetic poly[d(G-C)] and the poly(dC) strand of poly(dC).poly(dG) as well as single-stranded natural DNA that contains stretches of oligo(dC). The enhancing protein, a polypeptide of 65 kDa designated factor C, stimulates the copying of the two synthetic templates by Escherichia coli DNA polymerase I, Micrococcus luteus polymerase, and eukaryotic DNA polymerases alpha and beta, but not by avian myeloblastosis virus polymerase. Factor C, however, does not affect utilization by these polymerases of the poly(dG) strand of poly(dC).poly(dG), of poly(dC) primed by oligo(dG), or of poly(dA).poly(dT) and poly[d(A-T)]. With polymerase I, Michaelis constants (Km) of poly[d(G-C)] and of the poly(dC) strand of poly(dC).poly(dG) are decreased by factor C 37- and 4.7-fold, respectively, whereas maximum velocity (Vmax) remains unchanged. By contrast, neither the Km value of the poly(dG) strand of poly(dC).poly(dG) nor the Vmax value with this template is altered by factor C. Rates of copying of activated DNA, denatured DNA, or singly primed M13 DNA are not affected significantly by factor C. However, primer extension analysis of the copying of recombinant M13N4 DNA that contains runs of oligo(dC) within an inserted thymidine kinase gene shows that factor C increases processivity by specifically augmenting the efficiency at which polymerase I traverses the oligo(dC) stretches. Direct binding of factor C to denatured DNA is indicated by retention of the protein-DNA complex on columns of DEAE-cellulose. Binding of factor C to poly[d(G-C)] is demonstrated by the specific adsorption of the enhancing protein to columns of poly[d(G-C)]-Sepharose. We propose that by binding to poly[d(G-C)] and to poly(dC).poly(dG), factor C enables tighter binding of some DNA polymerases to these templates and facilitates enzymatic activity.
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PMID:Factor C from rabbit liver. A new poly(dC) and poly[d(G-C)] template-selective stimulatory protein of DNA polymerases. 292 91

Cytomegalovirus (CMV) DNA polymerase has immunologic specificity in relation to the virus specific polymerases of other herpesviruses. During the early phase of CMV infection in vitro, the virus DNA polymerase is rapidly induced. Due to the lack of virus specific thymidine kinase, cytomegalovirus is resistant to nucleosides which require herpesvirus thymidine kinases for activation. Cytomegalovirus DNA polymerase is therefore the known possible target for antiviral drugs. Several pyrophosphate analogs have been assayed for their inhibitory effects on this enzyme. The most active compound is phosphonoformate (PFA). PFA also effectively inhibits other partially purified herpesvirus DNA polymerases as well as the multiplication of all human herpesviruses, at concentrations which do not affect cellular DNA polymerases or normal cell growth. The mechanism of inhibition is a noncompetitive inhibition of CMV DNA polymerase activity with respect to the four deoxyribonucleoside triphosphates, and an uncompetitive inhibition with respect to the template used. In cell culture, PFA inhibits the formation of late CMV polypeptides, but not the synthesis of early CMV polypeptides. The CMV specific polymerase persists in the presence of PFA, as measured by immunological methods, and enzyme activity can be demonstrated after removal of PFA. The inhibition of CMV replication is reversible even after long exposure to PFA. Our interpretation is that the CMV genome is highly resistant to the cellular metabolism also in non-producing cells. A new rapid CMV neutralization test was established, based on the appearance of early CMV-induced antigens.
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PMID:Cytomegalovirus DNA polymerase inhibition and kinetics. 300 Jan 44

The antiherpetic agent 9-[(2,3-dihydroxy-1-propoxy)methyl]guanine (iNDG) is phosphorylated by HSV1 thymidine kinase, and its phosphorylated products inhibit DNA polymerase activity. iNDG exists in two enantiomeric forms, each with a primary and a secondary hydroxyl; thus, a number of possibilities for preferential phosphorylation exist, which were explored in this study. HSV1 thymidine kinase phosphorylates the primary hydroxyl of both the R and the S isomers of iNDG. This was established by comparison with analogues in which either the primary or the secondary hydroxyl was replaced by fluorine or hydrogen and also by a study of the NMR spectrum of the monophosphate. GMP kinase phosphorylates the R and the S monophosphates to the respective diphosphates. Further phosphorylation, however, is much more efficient with the S than with the R isomer. Furthermore, (S)-iNDG triphosphate is a more potent inhibitor of HSV1 DNA polymerase than (R)-iNDG triphosphate. These differences in the biochemical specificities of the two isomers account for the observed higher antiviral potency of (S)-iNDG as compared to that of (R)-iNDG.
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PMID:Enzymatic phosphorylation of the antiherpetic agent 9-[(2,3-dihydroxy-1-propoxy)methyl]guanine. 300 16

The activities of the purine acyclic nucleoside 9-(1,3-dihydroxy-2-propoxymethyl)guanine (DHPG) against two human and five animal strains of cytomegalovirus were compared with those of acyclovir. DHPG was significantly more active than acyclovir against all but one (mouse cytomegalovirus) of the strains tested, with 50% effective doses ranging from 5 to 13 microM, as determined by plaque reduction assays in human embryonic lung (MRC-5) and human embryonic tonsil cells. Both DHPG and acyclovir inhibited virus replication at concentrations considerably lower than those necessary to inhibit cell proliferation. In mode-of-action studies, the triphosphates of DHPG and acyclovir inhibited human cytomegalovirus DNA polymerase. DHPG phosphorylation to the active triphosphate was enhanced in infected cells; however, this enzymatic activity was unrelated to thymidine kinase. In animal studies, DHPG was slightly more effective than acyclovir in reducing mouse cytomegalovirus-induced mortality.
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PMID:Activity of 9-(1,3-dihydroxy-2-propoxymethyl)guanine compared with that of acyclovir against human, monkey, and rodent cytomegaloviruses. 301 Aug 40


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