Fluorescent RNA Aptamers as a Tool to Study RNA-Modifying Enzymes.

ABSTRACT: RNA-modifying enzymes are difficult to assay due to the absence of fluorometric substrates. Here we show that the Broccoli, a previously reported fluorescent RNA-dye complex, can be modified to contain N(6)-methyladenosine, a prevalent mRNA base modification. Methylated Broccoli is nonfluorescent but, upon demethylation by the RNA demethylases fat mass and obesity-associated protein (FTO) or ALKBH5, it binds and activates the fluorescence of its cognate fluorophore. We describe a high-throughput screen (HTS) for FTO inhibitors using the fluorogenic methylated Broccoli substrate HTS assay, which performs robustly with a Z' factor >0.8 in the LOPAC1280 library. This allowed the identification of novel high-affinity FTO inhibitors. Several of these compounds were selective for FTO over the re

SUBMITTER: Svensen N 

PROVIDER: S-EPMC4800036 | biostudies-literature | 2016 Mar

REPOSITORIES: biostudies-literature