Diastereomer separation of azobenzene-tethered oligodeoxyribonucleotides and determination of their absolute configurations by enzymatic digestion.

Two diastereomers were produced by the introduction of azobenzene-tethering prochiral linker (2,2-bis(hydroxymethyl)propionic acid) in the modified ODN, which had been used for the photoregulation of DNA functions. We found that this modified ODN with sequence 5'-...pNpXpN...-3' (p = phosphate; N = nucleoside; X = azobenzene residue) could be digested to pX (the phosphate at the 5' side of X was left) by an over excess of Phosphodiesterase I. By comparing the retention time of pX from the separated diastereomer with that of authentic R- or S-pX on chiral HPLC, absolute configuration could be easily determined.