NEW INSIGHTS INTO THE RESISTANCE OF ALPHA-OLIGONUCLEOTIDES TO NUCLEASES

Several studies have shown that alpha-oligonucleotides (alpha-ONs) are more resistant to degradation by nucleases than are beta-oligonucleot ides (beta-ONs), but a few exceptions have been reported. The present work indicates that the resistance of alpha-ONs to 3'-exonucleases con tained in calf or human sera strongly depends on their 3'-terminal seq uence. When the 2 last residues were ...A-G, ...C-A, or ...C-T, the de gradation rates in tissue culture media with fetal calf serum were sim ilar to those observed for beta-ONs, but stabilization factors up to 2 00 were observed when the 2 last residues were ...T-C, ...A-C, or ...C -C, and intermediate stabilization factors were found with ...G-A, ... T-A, or ...T-T terminal sequences. Other data confirm that alpha-ONs a re significantly more resistant than beta-ONs to purified 5'-exo- and endonucleases as well as to 3'-exonucleases contained in snake venom o r CEM cell lysates, but suggest that sequence specificity may also exi st in these media. These findings, which are consistent with literatur e data and explain the behavior of the aforementioned exceptions, also emphasize that the stability observed in the presence of purified nuc leases cannot be extrapolated to sera. Other results show that handlin g, heat inactivation, and storage conditions have little effect on the 3'-exonuclease activity of serum-containing media, but can completely modify the nuclease activities of cell extracts. This study, which wa s carried out by means of the accurate ''on-line ISRP cleaning'' HPLC technique, brings new insights to the general problem of oligonucleoti de stability.