Synthesis end properties of RNA analogs - oligoribonucleotide N3 '-> P5 ' phosphoramidates

The synthesis and characterization of RNA mimetics, uniformly modified olig oribonucleotide N3'-->P5' phosphoramidates containing all four natural base s (uracil, cytosine, adenine and guanine) as well as thymidine and 2,6-diam inopurine, are described, These RNA analogs contain N3'-->P5' phosphoramida te internucleotide linkages which replaced natural RNA O3'-->P5' phosphodie ster groups. These oligonucleotides were constructed from novel monomeric u nits (2'-t-butyldimethylsilyl)-3'-(monomethoxyltrityl)-amino-nucleoside-5'- phosphoramidites, the preparation of which is also presented. Several mixed base 9-13mer oligoribonucleotide phosphoramidates were synthesized with st ep-wise coupling yields of 96-98%, Thermal denaturation experiments demonst rated that ribo-N3'-->P5' phosphoramidates form stable duplexes with a comp lementary RNA strand. Thus, the melting temperature (T-m) of a duplex forme d by a 13mer ribo-N3'-->P5' phosphoramidate (84 degrees C) was higher than that observed for the isosequential natural RNA oligomer (64.0 degrees C), or for the 2'-deoxy-N3'-->P5' phosphoramidate counterpart (71.7 degrees C). Moreover, substitution of adenine by 2,6-diaminopurine in an oligoribophos phoramidate pentamer resulted in a very significant increase in the duplex melting temperature (similar to 7 degrees C per base substitution). The RNA phosphoramidates also showed similar rates of hydrolysis by both RNase A a nd RNase T-1 as compared to natural RNA oligomers, The data presented indic ate that this class of RNA analogs may be used as structural and functional RNA mimetics.