| Budget Amount *help |
¥3,600,000 (Direct Cost: ¥3,600,000)
Fiscal Year 2000: ¥2,000,000 (Direct Cost: ¥2,000,000)
Fiscal Year 1999: ¥1,600,000 (Direct Cost: ¥1,600,000)
|
|---|
| Research Abstract |
Amide-linked RNA derivatives and 2', 4'-aminomethyl-linked oligonucleotides were synthesized. Properties of these modified oligonucleotides were examined for the binding to the complimentary RNA in addition to the nuclease resistance.
1. Synthesis and properties of amide-linked RNA derivatives
The amide-linked uridylyl-uridine dinucleoside derivative was synthesized using 5'-N-monomethoxytrityl-5'-amino-3'-carboxymethyl-3', 5'-dideoxyuridine derivative and 5'-amino-5'deoxyuridine derivative. A phosphonium type of condensing reagent, PyBOP, was found to be effective for the rapid amide-bond formation. By means of 1H-NMR analysis, the ribose conformation of the upstream uridine unit was found to be fixed in the C3'-endo conformation. This condensing method was applied to the solid-phase synthesis of amide-linked RNA oligomers, U^*U^*U^*U^*U^*U (1), and CCCUUUU^*U^*U^*U (2)(^* : amide linkage). The UV-melting temperature of 2-RNA duplex was slightly lower to that of the unmodified duplex (Δ
… More
Tm-2℃). CD spectra analysis indicated that the 2-RNA duplex has A-form structure. These results suggest that the amide-linked RNA derivatives have RNA binding affinity in addition to the nuclease resistance.
2. Synthesis and properties of 2', 4'-aminomethyl-linked oligonucleotides
A 2', 4'-aminomethyl-linked uridylyl-thymidine dinucleoside phosphoramidite was synthesized in 11 steps from 1- (β-D-arabinofuranosyl) uracil with 5'-amino-5'-deoxythymidine. Oligonucleotides containing 2', 4'-aminomethyl linkages, dUaTUaTT (3) and dCCCTUaTTTTT (4) (a : 2', 4'-aminomethyl linkage), were synthesized using the UaT phosphoramidite by solid-phase synthesis. The oligomer 3 was digested by a 3'-exonuclease only at the 3'-terminal phosphodiester linkage. This result suggests that the 3'-terminal 2', 4'- aminomethyl internucleoside residue can prevent degradation of the oligonucleotide by 3-exonucleases. The oligomer 4 containing of a 2', 4'-aminomethyl linkage at the center of the strand showed relatively lower affinity to DNA and RNA.However, the CD spectra of 4-RNA duplex indicated A-form structure. This result suggests that the 2', 4'-aminomethyl linkage could incorporate into A-form structure without the global distortion. Less
|
