An unexpected methyl group migration during on-column Stille derivatization of RNA

Postsynthetic Stille cross-coupling for functionalization of oligonucleotides on solid support was applied on iodo modified RNA utilizing different protecting group strategies. As result, the otherwise very successful ACE [bis(acetoxyethyloxy)-methyl orthoester] chemistry was found to be limited since methylated side-products formed as was investigated via enzymatic degradation of RNA and various monomer model reactions. Enzymatic digestion of poly uridine sequences revealed presence of considerable amounts of N3-methylated uridine derivatives due to migration of methyl as phosphate protecting group used in ACE strategy. Monomer test reactions mimicking conditions on RNA clearly indicated an enhanced methylation effect correlated to the Stille coupling procedure.