The pH optimum of native uracil-DNA glycosylase of Archaeoglobus fulgidus compared to recombinant enzyme indicates adaption to cytosolic pH.

Knævelsrud, Ingeborg; Kazazic, Sabina; Birkeland, Nils-Kåre; Bjelland, Svein

doi:10.18388/abp.2014_1913

Cited by 3 publications

(1 citation statement)

References 27 publications

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“…fulgidus UDG has the optimal temperature of the enzyme activity at 80 o C (20). In this work, we demonstrated that Tba UDG194 has maximum cleavage efficiencies at >85 o C, which is close to optimal growth temperature of its host.…”

Section: Discussionsupporting

confidence: 54%

Characterization of a Family IV uracil DNA glycosylase from the hyperthermophilic euryarchaeon Thermococcus barophilus Ch5

Gan

Shi

et al. 2020

International Journal of Biological Macromolecules

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The hyperthermophilic euryarchaeon Thermococcus barophilus Ch5 encodes two uracil DNA glycosylases (UDGs): Tba UDG247 and Tba UDG194. In our previous publication, we revealed biochemical characterization of Tba UDG247. Herein, we characterized biochemically Tba UDG194, which is a member of Family IV UDG, demonstrating that this enzyme has similar efficiencies for cleaving uracil-containing ssDNA and dsDNA. Compared with Tba UDG247, Tba UDG194 exhibits different biochemical characteristics. At >85 o C, >90 cleavage percentage was observed, suggesting that Tba UDG194 can remove uracil from DNA at the close physiological temperature of its host. Thus, the enzyme has been currently the most thermophilic glycosylase among all the reported UDGs. Furthermore, the optimal pH of the enzyme activity was estimated to be 10, which is higher than that of Tba UDG247. Similar to Tba UDG247, Tba UDG194 activity is independent on a divalent metal ion. Mn 2+ , Zn 2+ and Cu 2+ display inhibitory effect on the enzyme activity at varied degreed whereas Mg 2+ and Ca 2+ have no detectable effect on the enzyme activity. In addition, Tba UDG194 is a salt-tolerant enzyme that retains compromised activity at 600 mM NaCl. Furthermore, Tba UDG 194 displays the following substrate preference: U≈U/ G>U/T≈U/C>U/A. The Arrhenius activation energy was etsimated to be 20.1 ± 3.4 kcal/mol, theoretically representing the energy barrier for uracil removal from DNA by Tba UDG194. Overall, our observations suggest that Tba UDG194 might be involved in removal of uracil in DNA in Thermococcus cells.

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Section: Discussionsupporting

confidence: 54%

Characterization of a Family IV uracil DNA glycosylase from the hyperthermophilic euryarchaeon Thermococcus barophilus Ch5

Gan

Shi

et al. 2020

International Journal of Biological Macromolecules

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Biochemical characterization and mutational studies of a thermostable uracil DNA glycosylase from the hyperthermophilic euryarchaeon Thermococcus barophilus Ch5

Shi

Gan

Jiang

et al. 2019

International Journal of Biological Macromolecules

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Uracil DNA glycosylases (UDGs) play an important role in removing uracil from DNA to initiate DNA base excision repair. Here, we first characterized biochemically a thermostable UDG from the hyperthermophilic euryarchaeon Thermococcus barophilus Ch5 (Tba UDG), and probed its mechanism by mutational analysis.The recombinant Tba UDG cleaves specifically uracil-containing ssDNA and dsDNA at 65 o C. The enzyme displays an optimal cleavage activity at 55-75 o C. Tba UDG cleaves DNA over a wide pH spectrum ranging from 4.0 to 9.0 with an optimal pH of 5.0-8.0. In addition, the Tba UDG activity is independent on a divalent metal ion; however, both Zn 2+ and Cu 2+ completely inhibits the enzyme activity. Furthermore, the Tba UDG activity is also inhibited by high NaCl concentration. Tba UDG removes uracil from DNA by the order: U≈U/G>U/T≈U/C>U/A. The mutational studies showed that both the E118A and N159A mutants completely abolish the cleavage activity and retain the compromised binding activity, suggesting that residues E118 and N159 in Tba UDG are important for uracil recognition and removal.Our work provides a basis for determining the role of Tba UDG in the base excision repair pathway for uracil repair in Thermococcus.

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Biochemical characterization and mechanistic insight of the family IV uracil DNA glycosylase from Sulfolobus islandicus REY15A

Lin

Dong

et al. 2023

International Journal of Biological Macromolecules

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The pH optimum of native uracil-DNA glycosylase of Archaeoglobus fulgidus compared to recombinant enzyme indicates adaption to cytosolic pH.

Cited by 3 publications

References 27 publications

Characterization of a Family IV uracil DNA glycosylase from the hyperthermophilic euryarchaeon Thermococcus barophilus Ch5

Characterization of a Family IV uracil DNA glycosylase from the hyperthermophilic euryarchaeon Thermococcus barophilus Ch5

Biochemical characterization and mutational studies of a thermostable uracil DNA glycosylase from the hyperthermophilic euryarchaeon Thermococcus barophilus Ch5

Biochemical characterization and mechanistic insight of the family IV uracil DNA glycosylase from Sulfolobus islandicus REY15A

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