Deoxyribonucleotide analogs as inhibitors and substrates of DNA polymerases

Wright, George E.; Brown, Neal C.

doi:10.1016/0163-7258(90)90066-b

Cited by 90 publications

(46 citation statements)

References 203 publications

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“…Pharmacologically, NRTIs have been divided into classes of mtDNA replication inhibitors according to the relative importance of DNA chain termination, or the internalization of the analog into nascent mtDNA and substitution for the natural base (Kakuda, 2000;Parker and Cheng, 1994;Wright and Brown, 1990). One class inhibits mtDNA replication in ways that resemble the action of fialuridine (FIAU).…”

Section: Nrti Pharmacological Classificationmentioning

confidence: 99%

Mitochondrial DNA Depletion, Oxidative Stress, and Mutation: Mechanisms 0f Dysfunction from Nucleoside Reverse Transcriptase Inhibitors

Lewis

Copeland

Day

2001

Laboratory Investigation

159

111

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Section: Nrti Pharmacological Classificationmentioning

confidence: 99%

Mitochondrial DNA Depletion, Oxidative Stress, and Mutation: Mechanisms 0f Dysfunction from Nucleoside Reverse Transcriptase Inhibitors

Lewis

Copeland

Day

2001

Laboratory Investigation

159

111

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“…Polymerization of these analogs frequently generates 3Ј termini that are poorly extended by DNA polymerases (15). The proofreading exonucleases associated with the mammalian DNA polymerases have been shown to remove some nucleotide analogs, but the efficiency of this excision is poor (16 -19).…”

mentioning

confidence: 99%

Identification and Expression of the TREX1 and TREX2 cDNA Sequences Encoding Mammalian 3′→5′ Exonucleases

Mazur

Perrino²

1999

Journal of Biological Chemistry

245

218

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The 335 exonucleases catalyze the excision of nucleoside monophosphates from the 3 termini of DNA. We have identified the cDNA sequences encoding two 335 exonucleases (TREX1 and TREX2) from mammalian cells. The TREX1 and TREX2 proteins are 304 and 236 amino acids in length, respectively. Analysis of the TREX1 and TREX2 sequences identifies three conserved motifs that likely generate the exonuclease active site in these enzymes. The specific amino acids in these three conserved motifs suggest that these mammalian exonucleases are most closely related to the proofreading exonucleases of the bacterial replicative DNA polymerases and the RNase T enzymes. Expression of TREX1 and TREX2 in Escherichia coli demonstrates that these recombinant proteins are active 335 exonucleases. The recombinant TREX1 protein was purified, and exonuclease activity was measured using single-stranded, partial duplex, and mispaired oligonucleotide DNA substrates. The greatest activity of the TREX1 protein was detected using a partial duplex DNA containing five mispaired nucleotides at the 3 terminus. No activity was detected using single-stranded RNA or an RNA-DNA partial duplex. Identification of the TREX1 and TREX2 cDNA sequences provides the genetic tools to investigate the physiological roles of these exonucleases in mammalian DNA replication, repair, and recombination pathways.The multistep processes of DNA replication, repair, and recombination in human cells often require the excision of nucleotides from the DNA 3Ј termini. For each cell division 4 billion nucleotides must be correctly replicated. The polymerization of incorrect or structurally modified nucleotides into DNA generates the 3Ј termini that block chain elongation by the DNA polymerases. Oxidative damage to DNA can result in fragmented nucleotides at the 3Ј termini that can not be elongated by DNA polymerases. Genetic recombination and mismatch repair pathways can require the removal of normal nucleotides from the 3Ј termini of DNA chains. Enzymes containing 3Ј35Ј exonuclease activity remove these mismatched, modified, fragmented, and normal nucleotides to generate the appropriate 3Ј termini for subsequent steps in the DNA metabolic pathways.Several 3Ј35Ј exonucleases have been described from a variety of animal cells (1-5). These exonucleases demonstrate similar biochemical properties, but the relationships between these enzymes are not known. Also, there are 3Ј35Ј exonucleases contained in the structural domains of mammalian DNA pols 1 ␦ (6), ⑀ (7), and ␥ (8). These proofreading 3Ј35Ј exonucleases excise incorrectly polymerized nucleotides during DNA synthesis. Thus, a variety of 3Ј35Ј exonucleases are present in mammalian cells. These exonucleases might function in multiple pathways to generate 3Ј termini that support further steps such as polymerization or ligation.Excision of incorrectly polymerized nucleotides by proofreading 3Ј35Ј exonucleases is an important mechanism to minimize errors during DNA synthesis. The polymerase-associated proofreading exonuclease was ...

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“…While several DNA repair polymerases (e.g., polymerases ␤, , , , and ) can be eliminated from consideration on the basis of their insensitivity to aphidicolin (23,34,37,59,64), PMEG and aphidicolin sensitivity do not serve to discriminate between damage repair and concatemer synthesis as potential roles for host DNA polymerases. The DNA polymerases that mediate host genome replication, polymerases ␣, ␦, and ⑀, are each sensitive to aphidicolin and PMEG (30,45,65) and are therefore prime candidates if host polymerases play a role in concatemer synthesis. However, polymerases ␦ and ⑀ are also involved in damage repair (54), and the DNA damage repair polymerases , , and are also sensitive to aphidicolin (46,55,57).…”

mentioning

confidence: 99%

Impact of 2-Bromo-5,6-Dichloro-1-β- d -Ribofuranosyl Benzimidazole Riboside and Inhibitors of DNA, RNA, and Protein Synthesis on Human Cytomegalovirus Genome Maturation

McVoy

Nixon

2005

J Virol

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Herpesvirus genome maturation is a complex process in which concatemeric DNA molecules are translocated into capsids and cleaved at specific sequences to produce encapsidated-unit genomes. Bacteriophage studies further suggest that important ancillary processes, such as RNA transcription and DNA synthesis, concerned with repeat duplication, recombination, branch resolution, or damage repair may also be involved with the genome maturation process. To gain insight into the biochemical activities needed for herpesvirus genome maturation, 2-bromo-5,6-dichloro-1-␤-D-ribofuranosyl benzimidazole riboside (BDCRB) was used to allow the accumulation of human cytomegalovirus concatemeric DNA while the formation of new genomes was being blocked. Genome formation was restored upon BDCRB removal, and addition of various inhibitors during this time window permitted evaluation of their effects on genome maturation. Inhibitors of protein synthesis, RNA transcription, and the viral DNA polymerase only modestly reduced genome formation, demonstrating that these activities are not required for genome maturation. In contrast, drugs that inhibit both viral and host DNA polymerases potently blocked genome formation. Radioisotope incorporation in the presence of a viral DNA polymerase inhibitor further suggested that significant host-mediated DNA synthesis occurs throughout the viral genome. These results indicate a role for host DNA polymerases in genome maturation and are consistent with a need for terminal repeat duplication, debranching, or damage repair concomitant with DNA packaging or cleavage. Similarities to previously reported effects of BDCRB on guinea pig cytomegalovirus were also noted; however, BDCRB induced low-level formation of a supergenomic species called monomer؉ DNA that is unique to human cytomegalovirus. Analysis of monomer؉ DNA suggested a model for its formation in which BDCRB permits limited packaging of concatemeric DNA but induces skipping of cleavage sites.The Herpesviridae family of viruses include several significant human pathogens, including herpes simplex virus type 1 (HSV-1) and -2, varicella-zoster virus, Epstein-Barr virus, Kaposi's sarcoma-associated herpesvirus, and human cytomegalovirus (HCMV). These viruses have large (130-to 235-kb) double-stranded linear DNA genomes that circularize shortly after infection (22,38,39,49). Viral DNA is replicated to form large concatemers of head-to-tail-linked genomes. In a process termed genome maturation, the concatemeric DNA is packaged into capsids and cleaved to produce encapsidated unit length genomes (6,8,27,36,38,49,52,66).Genome maturation is highly conserved among the herpesviruses, but little is known about the mechanisms or the machinery involved. Striking similarities to the DNA packaging mechanisms of certain large double-stranded DNA bacteriophages exist, in particular, , T3, T4, and T7 (14). In both bacteriophages and herpesviruses, procapsids self-assemble around a protein scaffold. A unique vertex of the procapsid contains portal proteins arra...

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Deoxyribonucleotide analogs as inhibitors and substrates of DNA polymerases

Cited by 90 publications

References 203 publications

Mitochondrial DNA Depletion, Oxidative Stress, and Mutation: Mechanisms 0f Dysfunction from Nucleoside Reverse Transcriptase Inhibitors

Mitochondrial DNA Depletion, Oxidative Stress, and Mutation: Mechanisms 0f Dysfunction from Nucleoside Reverse Transcriptase Inhibitors

Identification and Expression of the TREX1 and TREX2 cDNA Sequences Encoding Mammalian 3′→5′ Exonucleases

Impact of 2-Bromo-5,6-Dichloro-1-β- d -Ribofuranosyl Benzimidazole Riboside and Inhibitors of DNA, RNA, and Protein Synthesis on Human Cytomegalovirus Genome Maturation

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