1-beta-D-arabinofuranosyl-5-(E)-(4-azidostyryl)-uracil 5'-triphosphate: synthesis and utilization as a photoaffinity labeling reagent for DNA polymerase alpha from cherry salmon, Oncorhynchus masou

Biochem Biophys Res Commun. 1989 Jun 15;161(2):514-9. doi: 10.1016/0006-291x(89)92629-6.

Abstract

For developing a photoaffinity labeling reagent for DNA polymerase alpha, we synthesized 1-beta-D-arabinofuranosyl-5-(4-azidostyryl)-uracil 5'-triphosphate [5-(E)-(4-azidostyryl)-araUTP] bearing a photoreactive aryl azido group. This compound was easily decomposed by photolysis by light above 300 nm. This analogue strongly inhibited DNA polymerase alpha purified from cherry salmon, Oncorhynchus masou, testes; the mode of inhibition was mixed when the enzyme reaction was carried out under room light, and was competitive to dTTP in the dark. From the results of photoaffinity labeling experiments using an analogue containing [gamma-32P], it appeared that this analogue could bind to dTTP binding site of DNA polymerase alpha. Thus, this compound should be very useful for analysis of nucleotide binding sites of this enzyme.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Affinity Labels / chemical synthesis*
  • Animals
  • Azides / chemical synthesis*
  • Azides / pharmacology
  • DNA Polymerase I / antagonists & inhibitors
  • DNA Polymerase II / antagonists & inhibitors*
  • Kinetics
  • Male
  • Photochemistry
  • Salmon
  • Templates, Genetic
  • Testis / enzymology
  • Uracil Nucleotides / chemical synthesis*
  • Uridine Triphosphate / analogs & derivatives
  • Uridine Triphosphate / chemical synthesis*
  • Uridine Triphosphate / pharmacology

Substances

  • Affinity Labels
  • Azides
  • Uracil Nucleotides
  • 1-arabinofuranosyl-5-(4-azidostyryl)uracil 5'-triphosphate
  • DNA Polymerase I
  • DNA Polymerase II
  • Uridine Triphosphate