Synthesis of 2'-amine modified nucleosides and oligonucleotides to study RNA catalysis.

摘要

The specific role of the 2′-hydroxyl group in RNA catalysis has been traditionally addressed by studying reaction rates with 2′-deoxy, 2′-methoxy and 2 ′-amino analogues of the nucleosides, both at the nucleoside and oligonucleotide level. While these analogues did reveal the energetic contribution of the 2′-hydroxyl group toward catalysis, deciphering the exact role has been limited by simultaneous changes in steric bulk and electronic properties of the 2′-substituents. 2′-Methylamino analogues would be particularly helpful in this regard. Compared to the 2′-O-methyl analogue, the 2′-methylamino anlogue is sterically very similar but differs electronically in the ability of being a H-bond donor. Comparison of these two analogues might reveal the energetic cost of removing a H-bond donor at the 2′-position. On the other hand, comparison with the 2′-amino analogue might reveal the energetic cost of adding a methyl group at the 2′-position.; 2′-Amino-2′-deoxyguanosine has been previously incorporated into oligonucleotide, which involved HPLC purification of the free nucleoside enroute to the phosphoramidite. We have significantly simplified the procedure to obtain a phosphoramidite where the exocyclic amino group of the nucleobase was protected with a palmitoyl group. We showed that the phosphoramidite coupled efficiently in the solid phase synthesis and the palmitoyl group was removed quantitatively after oligonucleotide synthesis under the conditions of removal of other base-protecting groups.; 2′-Methylamino-2′-deoxyguanosine was synthesized as a probe to determine the role of the guanosine cofactor in the Tetrahymena group I reaction.; 2′-Methiaminopyrimidines have been incorporated into RNA. One such oligonucleotide (CCCUCU_2′-NHMe A) was used in the Tetrahymena group I reaction. Preliminary results demonstrate that the 2′-NHMe modification can be used to understand the 104 effect caused by modifying the 2′-OH (of the cleavage site uridine) to 2′ -OMe. We ascribe 102 fold to removal of the H-bond donor and 102 fold to installation of the CH ₃ group.