title: Benzimidazolium triflate-activated synthesis of (6-4) photoproduct-containing oligonucleotides and its application.
abstract: In the solid-phase synthesis of oligonucleotides containing the pyrimidine(6-4)pyrimidone photoproduct using a dinucleotide building block, considerable amounts of by-products were found as the chain length increased. The by-products were the major product when a 49mer was synthesized on a 40 nmol scale. It was assumed that these by-products were formed by the coupling of phosphoramidites with the N3 imino function of the 5' component of the (6-4) photoproduct. We examined imidazolium triflate and benzimidazolium triflate to find an alternative activator for DNA synthesis. Imidazolium triflate prevented by-product formation to some extent, but the coupling yields were low. Benzimidazolium triflate was comparable to tetrazole in coupling efficiency and reduced by-product formation to a great extent, without modification of the synthesizer program. The obtained 49mer was used to detect proteins that recognize UV-damaged DNA in HeLa cell extracts. Two major protein-DNA complexes were found when a 49mer duplex was used as probe, while a 30mer duplex failed to detect one of them. This application showed the usefulness of long chain 'damaged' oligonucleotides in biochemical studies.
mesh_terms: D001562:Benzimidazoles; D004260:DNA Repair; D004268:DNA-Binding Proteins / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D007093:Imidazoles; D009690:Nucleic Acid Conformation; D009838:Oligodeoxyribonucleotides / Q000096:biosynthesis* / Q000737:chemistry; D010777:Photochemistry; D011740:Pyrimidine Dimers / Q000737:chemistry / Q000378:metabolism / Q000528:radiation effects; D013777:Tetrazoles; D014466:Ultraviolet Rays