Characterisation of the Substrate Specificity of Homogeneous Vaccinia Virus Uracil-DNA Glycosylase

Overview

Journal Nucleic Acids Res

Publisher Oxford University Press

Specialty Biochemistry

Date 2003 Aug 9

PMID 12907738

Citations 22

Authors

Natale Scaramozzino

Guenhael Sanz

Jean Marc Crance

Murat Saparbaev

Robert Drillien

Jacques Laval

Bodil Kavli

Daniel Garin

Affiliations

Soon will be listed here.

Abstract

The decision to stop smallpox vaccination and the loss of specific immunity in a large proportion of the population could jeopardise world health due to the possibility of a natural or provoked re-emergence of smallpox. Therefore, it is mandatory to improve the current capability to prevent or treat such infections. The DNA repair protein uracil-DNA glycosylase (UNG) is one of the viral enzymes important for poxvirus pathogenesis. Consequently, the inhibition of UNG could be a rational strategy for the treatment of infections with poxviruses. In order to develop inhibitor assays for UNG, as a first step, we have characterised the recombinant vaccinia virus UNG (vUNG) and compared it with the human nuclear form (hUNG2) and catalytic fragment (hUNG) UNG. In contrast to hUNG2, vUNG is strongly inhibited in the presence of 7.5 mM MgCl(2). We have shown that highly purified vUNG is not inhibited by a specific uracil-DNA glycosylase inhibitor. Interestingly, both viral and human enzymes preferentially excise uracil when it is opposite to cytosine. The present study provides the basis for the design of specific inhibitors for vUNG.

Citing Articles

Unrepaired base excision repair intermediates in template DNA strands trigger replication fork collapse and PARP inhibitor sensitivity.

Serrano-Benitez A, Wells S, Drummond-Clarke L, Russo L, Thomas J, Leal G EMBO J. 2023; 42(18):e113190.

PMID: 37492888 PMC: 10505916. DOI: 10.15252/embj.2022113190.

Diatlova E, Mechetin G, Yudkina A, Zharkov V, Torgasheva N, Endutkin A Int J Mol Sci. 2023; 24(11).

PMID: 37298065 PMC: 10253040. DOI: 10.3390/ijms24119113.

A New Class of Uracil-DNA Glycosylase Inhibitors Active against Human and Vaccinia Virus Enzyme.

Grin I, Mechetin G, Kasymov R, Diatlova E, Yudkina A, Shchelkunov S Molecules. 2021; 26(21).

PMID: 34771075 PMC: 8587785. DOI: 10.3390/molecules26216668.

Displacement of Slow-Turnover DNA Glycosylases by Molecular Traffic on DNA.

Yudkina A, Endutkin A, Diatlova E, Moor N, Vokhtantsev I, Grin I Genes (Basel). 2020; 11(8).

PMID: 32751599 PMC: 7465369. DOI: 10.3390/genes11080866.

Aberrant repair initiated by the adenine-DNA glycosylase does not play a role in UV-induced mutagenesis in .

Zutterling C, Mursalimov A, Talhaoui I, Koshenov Z, Akishev Z, Bissenbaev A PeerJ. 2018; 6:e6029.

PMID: 30568855 PMC: 6286661. DOI: 10.7717/peerj.6029.

References

Holzer G, Falkner F . Construction of a vaccinia virus deficient in the essential DNA repair enzyme uracil DNA glycosylase by a complementing cell line. J Virol. 1997; 71(7):4997-5002. PMC: 191732. DOI: 10.1128/JVI.71.7.4997-5002.1997. View

Nilsen H, Otterlei M, Haug T, Solum K, Nagelhus T, Skorpen F . Nuclear and mitochondrial uracil-DNA glycosylases are generated by alternative splicing and transcription from different positions in the UNG gene. Nucleic Acids Res. 1997; 25(4):750-5. PMC: 146498. DOI: 10.1093/nar/25.4.750. View

Reddy S, Williams M, Cohen J . Expression of a uracil DNA glycosylase (UNG) inhibitor in mammalian cells: varicella-zoster virus can replicate in vitro in the absence of detectable UNG activity. Virology. 1998; 251(2):393-401. DOI: 10.1006/viro.1998.9428. View

Henderson D, INGLESBY T, Bartlett J, Ascher M, Eitzen E, Jahrling P . Smallpox as a biological weapon: medical and public health management. Working Group on Civilian Biodefense. JAMA. 1999; 281(22):2127-37. DOI: 10.1001/jama.281.22.2127. View

Otterlei M, Warbrick E, Nagelhus T, Haug T, Slupphaug G, Akbari M . Post-replicative base excision repair in replication foci. EMBO J. 1999; 18(13):3834-44. PMC: 1171460. DOI: 10.1093/emboj/18.13.3834. View

McCraith S, Holtzman T, Moss B, Fields S . Genome-wide analysis of vaccinia virus protein-protein interactions. Proc Natl Acad Sci U S A. 2000; 97(9):4879-84. PMC: 18326. DOI: 10.1073/pnas.080078197. View

Ishii K, Moss B . Role of vaccinia virus A20R protein in DNA replication: construction and characterization of temperature-sensitive mutants. J Virol. 2001; 75(4):1656-63. PMC: 114074. DOI: 10.1128/JVI.75.4.1656-1663.2001. View

Berche P . The threat of smallpox and bioterrorism. Trends Microbiol. 2001; 9(1):15-8. DOI: 10.1016/s0966-842x(00)01855-2. View

Klietmann W, Ruoff K . Bioterrorism: implications for the clinical microbiologist. Clin Microbiol Rev. 2001; 14(2):364-81. PMC: 88979. DOI: 10.1128/CMR.14.2.364-381.2001. View

10.

Nilsen H, Haushalter K, Robins P, Barnes D, Verdine G, Lindahl T . Excision of deaminated cytosine from the vertebrate genome: role of the SMUG1 uracil-DNA glycosylase. EMBO J. 2001; 20(15):4278-86. PMC: 149160. DOI: 10.1093/emboj/20.15.4278. View

11.

Lindahl T . Keynote: past, present, and future aspects of base excision repair. Prog Nucleic Acid Res Mol Biol. 2001; 68:xvii-xxx. DOI: 10.1016/s0079-6603(01)68084-x. View

12.

Klemperer N, McDonald W, Boyle K, Unger B, Traktman P . The A20R protein is a stoichiometric component of the processive form of vaccinia virus DNA polymerase. J Virol. 2001; 75(24):12298-307. PMC: 116126. DOI: 10.1128/JVI.75.24.12298-12307.2001. View

13.

Punjabi A, Boyle K, DeMasi J, Grubisha O, Unger B, Khanna M . Clustered charge-to-alanine mutagenesis of the vaccinia virus A20 gene: temperature-sensitive mutants have a DNA-minus phenotype and are defective in the production of processive DNA polymerase activity. J Virol. 2001; 75(24):12308-18. PMC: 116127. DOI: 10.1128/JVI.75.24.12308-12318.2001. View

14.

Kaplan E, Craft D, Wein L . Emergency response to a smallpox attack: the case for mass vaccination. Proc Natl Acad Sci U S A. 2002; 99(16):10935-40. PMC: 125076. DOI: 10.1073/pnas.162282799. View

15.

Kavli B, Sundheim O, Akbari M, Otterlei M, Nilsen H, Skorpen F . hUNG2 is the major repair enzyme for removal of uracil from U:A matches, U:G mismatches, and U in single-stranded DNA, with hSMUG1 as a broad specificity backup. J Biol Chem. 2002; 277(42):39926-36. DOI: 10.1074/jbc.M207107200. View

16.

Halloran M, Longini Jr I, Nizam A, Yang Y . Containing bioterrorist smallpox. Science. 2002; 298(5597):1428-32. DOI: 10.1126/science.1074674. View

17.

De Silva F, Moss B . Vaccinia virus uracil DNA glycosylase has an essential role in DNA synthesis that is independent of its glycosylase activity: catalytic site mutations reduce virulence but not virus replication in cultured cells. J Virol. 2002; 77(1):159-66. PMC: 140647. DOI: 10.1128/jvi.77.1.159-166.2003. View

18.

Lane J, Ruben F, Neff J, MILLAR J . Complications of smallpox vaccination, 1968. N Engl J Med. 1969; 281(22):1201-8. DOI: 10.1056/NEJM196911272812201. View

19.

Lindahl T, Nyberg B . Heat-induced deamination of cytosine residues in deoxyribonucleic acid. Biochemistry. 1974; 13(16):3405-10. DOI: 10.1021/bi00713a035. View

20.

Gupta P, Sirover M . Stimulation of the nuclear uracil DNA glycosylase in proliferating human fibroblasts. Cancer Res. 1981; 41(8):3133-6. View