A structure-guided strategy for modulating RNA-binding protein activity
RNA-binding proteins (RBPs) play critical roles in RNA stability, translation, and regulation. However, many RBPs remain challenging targets because they lack well-defined binding pockets for conventional small-molecule drugs.
MEX3C is one such challenging target. As an RNA-binding protein with E3 ubiquitin ligase activity, MEX3C promotes mRNA degradation and has been associated with tumor progression.
A recent study published in ACS Applied Bio Materials demonstrated a DNA aptamer-based strategy to selectively modulate MEX3C-mediated RNA regulation.
Engineering a High-Affinity DNA Aptamer
The researchers used a structure-guided Blocker-SELEX strategy to target the RNA-binding interface of the MEX3C KH1 domain.
After iterative screening and sequence optimization, they identified a 20-nt DNA aptamer, MRiApt, which showed:
- KD = 43.41 ± 2.64 nM for MEX3C-KH1
- IC50 = 10.19 ± 1.30 nM for blocking KH1–RNA binding
- High selectivity, with no significant effect on the homologous KH2 domain
Further optimization using phosphorothioate (PT) modification and stem-loop stabilization generated MRiApt-PT-stem, improving its performance to:
- KD = 5.26 ± 1.36 nM
- IC50 = 6.64 ± 0.31 nM
- Approximately 5.9 h serum half-life
- Efficient cellular uptake without transfection reagents
Demonstrating Activity in Cancer Cells
The optimized aptamer was then evaluated in cell-based assays.
MRiApt-PT-stem inhibited MEX3C-mediated degradation of HLA-A2 mRNA, increasing HLA-A2 mRNA levels by approximately 1.5-fold in HepG2 cells. The treatment also enhanced immune synapse formation between tumor cells and T cells, supporting the potential of MEX3C modulation in tumor immunity.
NMR analysis, molecular dynamics simulations, and alanine scanning further identified key residues involved in aptamer binding, providing structural insights into the MEX3C–aptamer interaction.
Figure: Screening of inhibitory DNA aptamers targeting the MEX3C-KH1 domain using the Blocker-SELEX strategy.
Supporting the Workflow with Yeasen Solutions
From recombinant protein preparation to cellular validation, reliable reagents were essential across multiple stages of the study.
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Product |
Cat. No. |
Application |
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20156ES50 |
Removing nucleic acid contamination during MEX3C-KH1 protein purification |
|
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DMEM, High Glucose |
41401ES76 |
HepG2 and HEK293T cell culture |
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RPMI 1640 Medium |
— |
Ramos cell culture |
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One-step RT-gDNA Digestion SuperMix |
11142ES60 |
Genomic DNA removal and reverse transcription in one step |
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11184ES08 |
qPCR quantification of HLA-A2 mRNA |
|
|
LysoTracker Lysosomal Probe |
40739ES50 |
Labeling lysosomes to trace aptamer intracellular localization |
Reference
Wei Y, et al. A DNA Aptamer as a Chemical Tool to Modulate MEX3C-Mediated mRNA Destabilization. ACS Applied Bio Materials. 2026. DOI: 10.1021/acsabm.5c02157.