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Split Hybridization Probe Utilizing a DNA Fluorescent Light-up Aptamer as a Signal Reporter for Sequence-Specific Nucleic Acid Analysis

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Structure-based investigation of fluorogenic Pepper aptamer.

Kaiyi Huang¹, Xianjun Chen^2,3, Chunyan Li¹

¹Life Sciences Institute, Zhejiang University, Hangzhou, China.

Nature Chemical Biology

|November 2, 2021

Summary

This summary is machine-generated.

Researchers elucidated the structure of Pepper aptamer, a novel RNA tag for live-cell imaging. This study reveals the molecular basis for its bright fluorescence, aiding future RNA visualization applications.

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Area of Science:

Molecular Biology
Biochemistry
Structural Biology

Background:

Pepper fluorescent RNAs offer bright, stable, multicolor imaging for diverse RNAs in live cells.
Understanding the molecular basis of Pepper's superior properties is crucial for optimizing its applications.

Purpose of the Study:

To determine the high-resolution structures of Pepper aptamer complexes with cognate fluorophores.
To investigate the molecular interactions governing Pepper aptamer's fluorescence activation mechanism.

Main Methods:

X-ray crystallography was employed to resolve the structures of Pepper aptamer-fluorophore complexes.
Structure-based mutational analysis was performed and evaluated using in vitro and live-cell assays.

Main Results:

The Pepper aptamer adopts a monomeric, non-G-quadruplex tuning-fork-like structure.
Fluorophores intercalate within the aptamer, positioned between a non-G-quadruplex base quadruple and a G·U wobble base pair.
Mutational analysis confirmed the structure-function relationship for fluorescence activation.

Conclusions:

This research provides a structural foundation for understanding Pepper aptamer's fluorescence mechanism.
The findings facilitate the rational design and improvement of RNA visualization tools.