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DNA Replication Fidelity with 8-Oxodeoxyguanosine Triphosphate
Abstract
Oxidative metabolism is known to generate mutagenic compounds within cells, among which is 8-oxodeoxyguanosine. Here the mutagenic potential of the triphosphate form of this base analog (8-O-dGTP) is investigated during replication in vitro of the lacZ alpha-complementation sequence in M13mp2 DNA. Adding 8-O-dGTP at equimolar concentration with the normal dNTPs to polymerization reactions decreases the fidelity of DNA synthesis by exonuclease-deficient Klenow, T4, and Thermus thermophilus DNA polymerases. Sequence analysis of mutants suggests that 8-O-dGMP is misincorporated opposite template adenines, yielding A-->C transversions. The degree of polymerase selectivity against this error is enzyme-dependent, with rates varying by > 25-fold. To determine if the A.8-O-dGMP mispair is proofread, a direct comparison of the fidelity of proofreading-proficient and proofreading-deficient Klenow and T4 DNA polymerases was made. Although the exonuclease activity of Klenow polymerase did not substantially reduce overall misincorporation of 8-O-dGMP, misincorporation was lower for the proofreading-proficient T4 enzyme as compared to its proofreading-deficient derivative. These data suggest that the A.8-O-dGMP mispair can be proofread. The mutagenic potential of 8-O-dGTP with eukaryotic systems was also examined. Misincorporation of 8-O-dGTP opposite adenine was observed during SV40 origin-dependent replication of double-stranded DNA in HeLa cell extracts. When present during replication at a concentration equal to the four normal dNTPs, 8-O-dGTP was at least 13-fold more mutagenic for A.T-->C.G transversions than was a 100-fold excess of normal dGTP.(ABSTRACT TRUNCATED AT 250 WORDS)
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“MtDNA oxidation, in turn, renders mtDNA more susceptible to mutation because oxidized bases are misread during replication and this leads to nucleotide substitutions [118–120].”
From Brain aging, Alzheimer's disease, and mitochondria (2011) · cites this paper as [120]
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Metadata
- Authors
- Pavlov, Y. I., Minnick, D. T., Izuta, S., & Kunkel, T. A.
- Affiliations
- National Institutes of Health (US)
- Venue
- Biochemistry
- Publisher
- American Chemical Society (ACS)
- Published
- 1994-04-19
- Volume
- 33
- Issue
- 15
- Pages
- 4695-4701
- ISSN
- 0006-2960 (ISSN-L)
- 1520-4995
- OpenAlex
- W1965072222
- Cited by
- 132
- References
- 43
- Citations by year
- 2026: 2 · 2025: 2 · 2024: 2 · 2021: 2 · 2020: 1 · 2019: 2 · 2018: 1 · 2017: 1 · 2016: 5 · 2015: 1 · 2014: 1 · 2013: 1
- FWCI
- 3.39
- Type
- journal article
- Language
- en
- Open access
- closed
- Locations
- Biochemistry (publishedVersion)
- PubMed (publishedVersion)
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › DNA Repair Mechanisms
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › DNA and Nucleic Acid Chemistry
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Cancer Research › Carcinogens and Genotoxicity Assessment
- Keywords
- 8-hydroxy-2'-deoxyguanosine, DNA replication fidelity, oxidative DNA damage, mutagenesis, misincorporation, DNA polymerase proofreading, exonuclease activity, Klenow fragment, T4 DNA polymerase, HeLa cell extracts, base analogs
- MeSH terms
- Base Composition, Deoxyguanine Nucleotides, DNA, DNA Polymerase I, DNA-Directed DNA Polymerase, DNA Replication, Escherichia coli, Glycosylation, Transfection, Mutagenesis, Thermus thermophilus, Bacteriophage T4
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