2'-Deoxyribonucleoside 5'-triphosphates bearing 4-phenyl and 4-pyrimidinyl imidazoles as DNA polymerase substrates

We developed a versatile access to a series of 4-substituted imidazole 2'-deoxynucleoside triphosphate bearing functionalized phenyl or pyrimidinyl rings. 4-Iodo-1H-imidazole was enzymatically converted into the corresponding 2'-deoxynucleoside, which was then chemically derived into its 5'-triphosphate, followed by 4-arylation via Suzuki-Miyaura coupling using (hetero)arylboronic acids. Both KF (exo-) and Deep Vent (exo-) DNA polymerases incorporated these modified nucleotides in primer-extension assays, adenine being the preferred pairing partner in the template. The 4-(3-aminophenyl)imidazole derivative (3APh) was the most efficiently inserted opposite A by KF (exo-) with only a 37-fold lower efficiency (Vmax/KM) than that of the correct dTTP. No further extension occurred after the incorporation of a single aryl-imidazole nucleotide. Interestingly, the aryl-imidazole dNTPs were found to undergo successive incorporation by calf thymus terminal deoxynucleotidyl transferase with different tailing efficiencies among this series and with a marked preference for 2APyr polymerization.