Chemical Synthesis of Oligoribonucleotide (ASL of tRNALys T. brucei) Containing a Recently Discovered Cyclic Form of 2-Methylthio-N6-threonylcarbamoyladenosine (ms2ct6A)

The synthesis of the protected form of 2-methylthio-N⁶-threonylcarbamoyl adenosine ( ms²t⁶A) was developed starting from adenosine or guanosine by using the optimized carbamate method and, for the first time, an isocyanate route. The hypermodified nucleoside was subsequently transformed into the protected ms²t⁶A -phosphoramidite monomer and used in a large-scale synthesis of the precursor 17nt ms²t⁶A -oligonucleotide (the anticodon stem and loop fragment of tRNA^Lys from T. brucei). Finally, stereochemically secure ms²t⁶A → ms²ct⁶A cyclization at the oligonucleotide level efficiently afforded a tRNA fragment bearing the ms²ct⁶A unit. The applied post-synthetic approach provides two sequentially homologous ms²t⁶A - and ms²ct⁶A -oligonucleotides that are suitable for further comparative structure–activity relationship studies.