A New Variant of Emissive RNA Alphabets

A new fluorescent ribonucleoside alphabet (^mth N) consisting of pyrimidine and purine analogues, all derived from methylthieno[3,4-d]pyrimidine as the heterocyclic core, is described. Large bathochromic shifts and high microenvironmental susceptibility of their emission relative to previous alphabets derived from thieno[3,4-d]pyrimidine (^th N) and isothiazole[4,3-d]pyrimidine (^tz N) scaffolds are observed. Subjecting the purine analogues to adenosine deaminase, guanine deaminase and T7 RNA polymerase indicate that, while varying, all but one enzyme tolerate the corresponding ^mth N/^mth NTP substrates. The robust emission quantum yields, high photophysical responsiveness and enzymatic accommodation suggest that the ^mth N alphabet is a biophysically viable tool and can be used to probe the tolerance of nucleoside/tide-processing enzymes to structural perturbations of their substrates.