SEARCH

SEARCH BY CITATION

Keywords:

  • Antisense agents;
  • Electron transfer;
  • Oligonucleotides;
  • Photochemistry;
  • Ruthenium

Abstract

The special photoreactivity of Ru-TAP (TAP = 1,4,5,8-tetraazaphenanthrene, phen = 1,10-phenanthroline) complexes anchored to G-containing oligodeoxyribonucleotide (ODN) probes prompted us to investigate thoroughly the parameters that influence their selectivity in two competing processes: the intermolecular photoreaction with the target complementary sequence, leading to crosslinking at G sites of the target, and the intramolecular photoreaction leading to a self-inhibition also called the Seppuku process. It is indeed particularly important to favour self-inhibition in the presence of mismatch-containing target sequences. It was found that any parameter reinforcing the association of the Ru-ODN conjugate with a sequence possessing impaired complementarity, such as a low temperature (22 °C), high sodium chloride concentration ([NaCl] = 150 mM), and the presence of an intercalating ligand in the Ru complex, led to increases in undesired photocrosslinking with the wrong target (i.e. that sequence bearing impaired complementarity). In contrast, less favorable conditions for hybridization (T = 33 °C, [NaCl] = 50 mM, a non-intercalating complex), led to a rather good selectivity; up to a 3:1 self-inhibition/photocrosslinking ratio was obtained with a target sequence containing only two mismatches. It is shown that this selectivity as well as the selectivity in the photocrosslinking sites, result from subtle changes in the probe and target interactions. These findings provide the basis for further improvements in selective DNA photoreagents (seppuku conjugates) for cancer research and elaborate the guidelines for selecting appropriate target sequences in a gene to be silenced.