Thioester Electrophiles as a New Class of SHAPE Probes

ABSTRACT

RNA is a multifunctional molecule capable of regulating gene expression, in large part because it can form a variety of RNA secondary and tertiary structures. The emergence of RNA viruses like SARS-CoV-2 emphasizes the need to accelerate our understanding of how viral RNA structure dictates its function. One approach to map RNA secondary structure, called Selective 2'-OH Acylation Analyzed by Primer Extension (SHAPE), utilizes select electrophiles that unbiasedly modifies the 2'-hydroxyl of riboses in unpaired nucleotides, forming adducts that are detected through a variety of sequencing methods. While SHAPE is widely utilized, most existing SHAPE reagents suffer from several drawbacks 1) poor water solubility;2) limited commercial availability;and 3) they function optimally when freshly synthesized, requiring synthetic organic expertise. To overcome these obstacles, our goal is to develop a userfriendly SHAPE reagent kit that provides highly reactive, soluble SHAPE reagents capable of probing RNA structure in vitro as well as in vivo. We present our investigations on developing thioester electrophiles as a new class of SHAPE reagents. Our reagent is prepared by mixing two stable components to generate the reactive thioester electrophile in situ. We report our preliminary results in model systems and the scope to expand the library of our reagents.Copyright © 2023 The American Society for Biochemistry and Molecular Biology, Inc.