Rational development of FMN-based orthogonal riboswitch that functions in response to specific non-cognate ligand

Divya Ojha and Ambadas B. Rode ()
Additional contact information
Divya Ojha: Faridabad-Gurugram Expressway
Ambadas B. Rode: Faridabad-Gurugram Expressway

Nature Communications, 2025, vol. 16, issue 1, 1-16

Abstract: Abstract Re-engineering natural riboswitches into orthogonal RNA switches by making them functional in response to exogenous ligands but unresponsive to endogenous cognate ligands is a promising yet less explored strategy for developing gene regulatory tools. Herein, we rationally engineer the aptamer domain of one of the largest and biotechnologically relevant flavin mononucleotide (FMN) riboswitch class, which specifically binds to synthetic ligands with a high binding affinity (KD = ~ 54-75 nM) and regulates gene expression in vitro, in prokaryotic, and eukaryotic system, while being unresponsive to FMN. To develop the orthogonal aptamers, we rationally alter key tertiary interactions, such as A/G minor motifs and base triples located in the periphery of the FMN binding pocket. The biophysical and structural probing analysis of the orthogonal aptamer and synthetic ligand complex shows binding mediated by favorable enthalpic and unfavorable entropic contributions. Our rational design approach, coupled with the adaptability to FMN aptamers derived from diverse bacterial strains, suggests the broad applicability of this strategy to numerous FMN riboswitches, each possessing a unique expression platform. This will greatly expand the current repertoire of synthetic riboswitches available for biomedical applications.

Date: 2025
References: Add references at CitEc
Citations:

Downloads: (external link)
https://www.nature.com/articles/s41467-025-60722-4 Abstract (text/html)

Related works:
This item may be available elsewhere in EconPapers: Search for items with the same title.

Export reference: BibTeX RIS (EndNote, ProCite, RefMan) HTML/Text

Persistent link: https://EconPapers.repec.org/RePEc:nat:natcom:v:16:y:2025:i:1:d:10.1038_s41467-025-60722-4

Ordering information: This journal article can be ordered from
https://www.nature.com/ncomms/

DOI: 10.1038/s41467-025-60722-4

Access Statistics for this article

Nature Communications is currently edited by Nathalie Le Bot, Enda Bergin and Fiona Gillespie

More articles in Nature Communications from Nature
Bibliographic data for series maintained by Sonal Shukla () and Springer Nature Abstracting and Indexing ().