Pepper RNA variants reveal decoupling of HBC530 binding thermodynamics and fluorescence activation.

ABSTRACT: Fluorogenic RNA aptamers have emerged as powerful tools for live-cell imaging and synthetic biology applications due to their ability to activate fluorescence upon ligand binding. However, the sequence-structure-function relationships governing ligand recognition and fluorescence activation remain poorly understood, limiting rational aptamer design. The Pepper aptamer binds HBC530 with nanomolar affinity in a magnesium-dependent manner, producing bright fluorescence suitable for cellular applications. Here, we generated a library of 53 Pepper variants containing substitutions, insertions, and/or deletions to quantitatively evaluate the contributions of individual nucleotides to HBC530 binding affinity, magnesium binding affinity, and fluorescence intensity. Our results reveal that the corr

SUBMITTER: Aidoo EK 

PROVIDER: S-EPMC12262465 | biostudies-literature | 2025 Jun

REPOSITORIES: biostudies-literature