NOVEL SOLID-PHASE SYNTHESIS OF BRANCHED OLIGORIBONUCLEOTIDES, INCLUDING A SUBSTRATE FOR THE RNA DEBRANCHING ENZYME

An effective new route for synthesizing branched oligoribonucleotides in the solid phase in the 5' to 3' direction has been developed. This required the synthesis of reversed monomers, viz. protected nucleoside 5'-phosphoramidites bearing 2'-O-Fpmp and 3'-O-pixyl protecting group s as well as special branch-point monomers, viz. protected nucleoside 5'-phosphoramidites bearing either 2',3'-O-dipixyl protection in the c ase of adenosine, cytidine and uridine, or 2',3'-O-dilaevulinyl protec tion in the case of guanosine. These monomers are assembled on commerc ial synthesizers into branched oligoribonucleotides in high yield, the crude products are readily purified by reversed-phase HPLC whilst sti ll partially protected, and the fully deprotected products are conveni ently analysed by electrospray mass spectrometry. Moreover, the branch ed oligoribonucleotides can be recognised and cleaved by a specific 2' -5' phosphodiesterase present in mammalian cell nuclei. We expect that this will prove valuable for future biochemical and biological studie s on the properties of branched RNA molecules and the protein factors and enzymes that interact with branched RNA substrates.