Structural Basis for Fluorescence Activation by Pepper RNA.

Publication Type:

Journal Article


ACS Chem Biol, Volume 17, Issue 7, p.1866-1875 (2022)


Binding Sites, Fluorescence, G-Quadruplexes, Ligands, Nucleic Acid Conformation, RNA


<p>Pepper is a fluorogenic RNA aptamer tag that binds to a variety of benzylidene-cyanophenyl (HBC) derivatives with tight affinity and activates their fluorescence. To investigate how Pepper RNA folds to create a binding site for HBC, we used antibody-assisted crystallography to determine the structures of Pepper bound to HBC530 and HBC599 to 2.3 and 2.7 Å resolutions, respectively. The structural data show that Pepper folds into an elongated structure and organizes nucleotides within an internal bulge to create the ligand binding site, assisted by an out-of-plane platform created by tertiary interactions with an adjacent bulge. As predicted from a lack of K dependence, Pepper does not use a G-quadruplex to form a binding pocket for HBC. Instead, Pepper uses a unique base-quadruple&middot;base-triple stack to sandwich the ligand with a U&middot;G wobble pair. Site-bound Mg ions support ligand binding structurally and energetically. This research provides insight into the structural features that allow the Pepper aptamer to bind HBC and show how Pepper&#39;s function may expand to allow the in vivo detection of other small molecules and metals.</p>

7SZU, 7U0Y