Repurposing an adenine riboswitch into a fluorogenic imaging and sensing tag.
| Publication Type | Academic Article |
| Authors | Dey S, Filonov G, Olarerin-George A, Jackson B, Finley L, Jaffrey S |
| Journal | Nat Chem Biol |
| Volume | 18 |
| Issue | 2 |
| Pagination | 180-190 |
| Date Published | 12/22/2021 |
| ISSN | 1552-4469 |
| Keywords | Aptamers, Nucleotide, Biosensing Techniques, Optical Imaging |
| Abstract | Fluorogenic RNA aptamers are used to genetically encode fluorescent RNA and to construct RNA-based metabolite sensors. Unlike naturally occurring aptamers that efficiently fold and undergo metabolite-induced conformational changes, fluorogenic aptamers can exhibit poor folding, which limits their cellular fluorescence. To overcome this, we evolved a naturally occurring well-folded adenine riboswitch into a fluorogenic aptamer. We generated a library of roughly 1015 adenine aptamer-like RNAs in which the adenine-binding pocket was randomized for both size and sequence, and selected Squash, which binds and activates the fluorescence of green fluorescent protein-like fluorophores. Squash exhibits markedly improved in-cell folding and highly efficient metabolite-dependent folding when fused to a S-adenosylmethionine (SAM)-binding aptamer. A Squash-based ratiometric sensor achieved quantitative SAM measurements, revealed cell-to-cell heterogeneity in SAM levels and revealed metabolic origins of SAM. These studies show that the efficient folding of naturally occurring aptamers can be exploited to engineer well-folded cell-compatible fluorogenic aptamers and devices. |
| DOI | 10.1038/s41589-021-00925-0 |
| PubMed ID | 34937909 |
| PubMed Central ID | PMC8967656 |