A new one-step method for the preparation of 3',5'-bisphosphates of acid-labile deoxynucleosides

A method is described for preparing 3',5'-bisphosphates of labile deoxynucleosides. Under strictly anhydrous conditions, pyrophosphoryl tetrakistriazole apparently forms a ring structure bridging the 3'- and 5'-hydroxyl groups of deoxynucleosides, since upon the addition of water the ring opens and the 3',5'-bisphosphate is formed. Due to the presence of triethylamine no acid is generated at any time so that the entire procedure is in neutral solution. The bisphosphates of N2,3-ethenodeoxyguanosine, O2-ethyldeoxythymidine, and O4-methyldeoxythymidine, all of which are acid-labile, were prepared in good yield without degradation. Other modified bisphosphates prepared include O6-benzyldeoxyguanosine and 1,N6-ethenodeoxyadenosine, as well as those of unmodified deoxyguanosine and thymidine. Characterization was by 31P NMR and UV spectroscopy. Both 5'p(dT)p3' and 5'p(dG)p3' were substrates for RNA ligase, further proving the structure of the phosphorylated compounds.