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Synthesis and characterization of oligonucleotides containing 2 '-fluorinated thymidine glycol as inhibitors of the endonuclease iii reaction

Academic Article
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Overview

authors

  • Doi, Y.
  • Katafuchi, A.
  • Fujiwara, Y.
  • Hitomi, K.
  • Tainer, John
  • Ide, H.
  • Iwai, S.

publication date

  • 2006

journal

  • Nucleic Acids Research  Journal

abstract

  • Endonuclease III (Endo III) is a base excision repair enzyme that recognizes oxidized pyrimidine bases including thymine glycol. This enzyme is a glycosylase/lyase and forms a Schiff base-type intermediate with the substrate after the damaged base is removed. To investigate the mechanism of its substrate recognition by X-ray crystallography, we have synthesized oligonucleotides containing 2'-fluorothymidine glycol, expecting that the electron-withdrawing fluorine atom at the 2' position would stabilize the covalent intermediate, as observed for T4 endonuclease V (Endo V) in our previous study. Oxidation of 5'- and 3'-protected 2'-fluorothymidine with OsO4 produced two isomers of thymine glycol. Their configurations were determined by NMR spectroscopy after protection of the hydroxyl functions. The ratio of (5R,6S) and (5S,6R) isomers was 3:1, whereas this ratio was 6:1 in the case of the unmodified sugar. Both of the thymidine glycol isomers were converted to the corresponding phosphoramidite building blocks and were incorporated into oligonucleotides. When the duplexes containing 2'-fluorinated 5R- or 5S-thymidine glycol were treated with Escherichia coli endo III, no stabilized covalent intermediate was observed regardless of the stereochemistry at C5. The 5S isomer was found to form an enzyme-DNA complex, but the incision was inhibited probably by the fluorine-induced stabilization of the glycosidic bond.

subject areas

  • Deoxyribonuclease (Pyrimidine Dimer)
  • Enzyme Inhibitors
  • Escherichia coli Proteins
  • Isomerism
  • Oligonucleotides
  • Organophosphorus Compounds
  • Substrate Specificity
  • Thymidine
  • Viral Proteins
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Identity

PubMed Central ID

  • PMC1409675

International Standard Serial Number (ISSN)

  • 0305-1048

Digital Object Identifier (DOI)

  • 10.1093/nar/gkl061

PubMed ID

  • 16547199
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Additional Document Info

start page

  • 1540

end page

  • 1551

volume

  • 34

issue

  • 5

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