1332478-75-7Relevant academic research and scientific papers
Fluorophore-Promoted RNA Folding and Photostability Enables Imaging of Single Broccoli-Tagged mRNAs in Live Mammalian Cells
Jaffrey, Samie R.,Kim, Hyaeyeong,Li, Xing,Litke, Jacob L.,Wu, Jiahui
, p. 4511 - 4518 (2020)
Spinach and Broccoli are fluorogenic RNA aptamers that bind DFHBI, a mimic of the chromophore in green fluorescent protein, and activate its fluorescence. Spinach/Broccoli-DFHBI complexes exhibit high fluorescence in vitro, but they exhibit lower fluorescence in mammalian cells. Here, computational screening was used to identify BI, a DFHBI derivative that binds Broccoli with higher affinity and leads to markedly higher fluorescence in cells compared to previous ligands. BI prevents thermal unfolding of Broccoli at 37 °C, leading to more folded Broccoli and thus more fluorescent Broccoli-BI complexes in cells. Broccoli-BI complexes are more photostable owing to impaired photoisomerization and rapid unbinding of photoisomerized cis-BI. These properties enable single mRNA containing 24 Broccoli aptamers to be imaged in live mammalian cells treated with BI. Small molecule ligands can thus promote RNA folding in cells, and thus allow single mRNA imaging with fluorogenic aptamers.
Bifacial peptide nucleic acid as an allosteric switch for aptamer and ribozyme function
Xia, Xin,Piao, Xijun,Bong, Dennis
supporting information, p. 7265 - 7268 (2014/06/09)
We demonstrate herein that bifacial peptide nucleic acid (bPNA) hybrid triplexes functionally substitute for duplex DNA or RNA. Structure-function loss in three non-coding nucleic acids was inflicted by replacement of a duplex stem with unstructured oligo-T/U strands, which are bPNA binding sites. Functional rescue was observed on refolding of the oligo-T/U strands into bPNA triplex hybrid stems. Bifacial PNA binding was thus used to allosterically switch-on protein and small-molecule binding in DNA and RNA aptamers, as well as catalytic bond scission in a ribozyme. Duplex stems that support the catalytic site of a minimal type I hammerhead ribozyme were replaced with oligo-U loops, severely crippling or ablating the native RNA splicing function. Refolding of the U-loops into bPNA triplex stems completely restored splicing function in the hybrid system. These studies indicate that bPNA may have general utility as an allosteric trigger for a wide range of functions in non-coding nucleic acids.
Plug-and-play fluorophores extend the spectral properties of spinach
Song, Wenjiao,Strack, Rita L.,Svensen, Nina,Jaffrey, Samie R.
supporting information, p. 1198 - 1201 (2014/02/14)
Spinach and Spinach2 are RNA aptamers that can be used for the genetic encoding of fluorescent RNA. Spinach2 binds and activates the fluorescence of (Z)-4-(3,5-difluoro-4-hydroxybenzylidene)-1,2-dimethyl-1H-imidazol-5(4H)-one (DFHBI), allowing the dynamic localizations of Spinach2-tagged RNAs to be imaged in live cells. The spectral properties of Spinach2 are limited by DFHBI, which produces fluorescence that is bluish-green and is not optimized for filters commonly used in fluorescence microscopes. Here we characterize the structural features that are required for fluorophore binding to Spinach2 and describe novel fluorophores that bind and are switched to a fluorescent state by Spinach2. These diverse Spinach2-fluorophore complexes exhibit fluorescence that is more compatible with existing microscopy filter sets and allows Spinach2-tagged constructs to be imaged with either GFP or YFP filter cubes. Thus, these plug-and-play fluorophores allow the spectral properties of Spinach2 to be altered on the basis of the specific spectral needs of the experiment.
Understanding the photophysics of the Spinach-DFHBI RNA aptamer-fluorogen complex to improve live-cell RNA imaging
Han, Kyu Young,Leslie, Benjamin J.,Fei, Jingyi,Zhang, Jichuan,Ha, Taekjip
, p. 19033 - 19038 (2014/01/06)
The use of aptamer-fluorogen complexes is an emerging strategy for RNA imaging. Despite its promise for cellular imaging and sensing, the low fluorescence intensity of the Spinach-DFHBI RNA aptamer-fluorogen complex hampers its utility in quantitative liv
