| Research Abstract |
In the antisense technique, one of the most promising diagnostic methods for severe viral diseases, . phosphorothioate oligodeoxynucleotides (S-oligos) which has a thiophosphate linkage have been investigated exclusively. To apply this method in clinic, however, there has been a crucial unsolved problem that one should identify and prepare the truly effective specific structure among numbers of stereoisomeric S-oligos arising from the stereoisomerism of each thiophosphate linkage, and in the present study, therefore, we have performed following investigation s to overcome the above problem. (1) We have carried out 2D-NMR analysis combined with molecular dynamic calculation on the relationship between the thiophosphate Rp and Sp configurations and the S-oligo RNA/DNA duplex structure. It has been elucidated that the structure of the duplexes with RNA is similar with each other as well as with that of wild type DNA-RNA duplex, although Rp linkage produced higher Tm, indicative that the d
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uplex stability is dependent such a factor other than the duplex structure, possibly as extent of hydration. On the other hand, difference of the duplex structure was found to be responsible for S-oligo-DNA hybrid stability. These new findings indicate that the, suitable configuration depends on the target, and that the mechanism of this rule also on it. (2). Nonspecific antiviral activity of S-oligo polyC has also been studied, and found to be due to its unique four-stranded solution structure, i-motif, whose stability is dependent on its Rp or Sp configuration. Also, a possible target gene for S-oligo polyC, telomere C-repeat, has been investigated in terms of the structure, and it has been found that there are isomeric C repeat tetrads. This new discovery may give suggestion not only to understand whole mechanism for nonspecific activity of S-oligo but biological functions of telomere topologies in relation to cancer and aging. (3) Stereoselective synthesis of S-oligos, another unsolved essential subject, has been studied and use of m-chlorocresol as a leaving group in the phosphite method was found to lead to high stereoselectivity and high yield of transesterification in the basic conditions. Less
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