Aqueous heck cross-coupling preparation of acrylate-modified nucleotides and nucleoside triphosphates for polymerase synthesis of acrylate-labeled DNA

摘要

Aqueous-phase Heck coupling methodology was developed for direct attachment of butyl acrylate to 5-iodoracil, 5-iodocytosine, 7-iodo-7-deazaadenine, and 7-iodo-7-deazaguanine 2′-deoxyribonucleoside 5′-O-monophosphates (dNMPs) and 5′-O-triphosphates (dNTPs) and compared with the classical approach of phosphorylation of the corresponding modified nucleosides. The 7-substituted 7-deazapurine nucleotides (dA~(BA)MP, dA~(BA)TP, dG~(BA)MP, and dG~(BA)TP) were prepared by the direct Heck coupling of nucleotides in good yields (35-55%), whereas the pyrimidine nucleotides reacted poorly and the corresponding BA-modified dNTPs were prepared by triphosphorylation of the modified nucleosides. The acrylate-modified dN~(BA)TPs (N = A, C, and U) were good substrates for DNA polymerases and were used for enzymatic synthesis of acrylate-modified DNA by primer extension, whereas dG~(BA)TP was an inhibitor of polymerases. The butyl acrylate group was found to be a useful redox label giving a strong reduction peak at -1.3 to -1.4 V in cyclic voltammetry.