Synthesis of oligonucleotides containing the (4R) and (4S) diastereoisomers of 4,8-dihydro-4-hydroxy-8-oxo-2 '-deoxyguanosine

The insertion of the (4R) and (4S) diastereoisomers of 4,8-dihydro-4-hydrox y-8-oxo-2'-deoxyguanosin the two main singlet oxygen oxidation products of 2'-deoxyguanosine, (4R)-4-OH-8-oxodGuo (3a) and (4S)-4-OH-8-oxodGuo (3b), i nto oligonucleotides has been achieved by means of phosphoramidite chemistr y using the solid-phase synthesis approach. The synthesis of the phosphoram idite synthons 8a and 8b required 6 steps from 2'-deoxyguanosine and involv ed the simultaneous protection of the additional tertiary hydroxy (4-OH) an d N-2-amino groups of the modified base by acetylation. The modified phosph oramidites 8a, b were efficiently incorporated into several oligonucleotide s (3-mer, 9-mer, 14-mer, and 22-mer). The presence and the integrity of the 3a and 3b in the synthetic oligomers was confirmed by electrospray ionizat ion mass spectrometry, together with HPLC and MALDI-TOF mass-spectrometric analyses of enzymatic digestions. The use of various enzymes, including end onucleases (nuclease P-1) and exonucleases (snake venom phosphodiesterase a nd calf spleen phosphodiesterase), clearly shows that the enzymatic hydroly sis of phosphodiester bonds between 3a,b and normal 2'-deoxyribonucleosides is inhibited.