Selection of RNA‐Cleaving TNA Enzymes for Cellular Mg2+ Imaging

Live-cell imaging: New RNA-cleaving TNA enzymes (TNAzymes) have been identified by in vitro selection. TNAzyme 17–22 catalyzes RNA cleavage with a k _cat of 0.017 min⁻¹ in the presence of Mg²⁺. The TNAzyme-based fluorescent sensor has a limit of detection of 0.35 mM and enables Mg²⁺ imaging in living cells.

Abstract

Catalytic DNA-based fluorescent sensors have enabled cellular imaging of metal ions such as Mg²⁺. However, natural DNA is prone to nuclease-mediated degradation. Here, we report the in vitro selection of threose nucleic acid enzymes (TNAzymes) with RNA endonuclease activities. One such TNAzyme, T17–22, catalyzes a site-specific RNA cleavage reaction with a k _cat of 0.017 min⁻¹ and K _M of 675 nM. A fluorescent sensor based on T17–22 responds to an increasing concentration of Mg²⁺ with a limit of detection at 0.35 mM. This TNAzyme-based sensor also allows cellular imaging of Mg²⁺. This work presents the first proof-of-concept demonstration of using a TNA catalyst in cellular metal ion imaging.

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