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Published on: April 26, 2017
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Rational design yields RNA-binding zinc finger domains with altered sequence specificity
Qishan Liang1,2,3,4, Joy S Xiang2, Gene W Yeo5,3,4
1Department of Chemistry and Biochemistry, UC San Diego, La Jolla, California 92093, USA.
RNA (New York, N.Y.)
|December 10, 2024
Summary
Researchers engineered RNA-binding zinc fingers (ZnFs) to alter RNA sequence specificity. This work advances programmable RNA targeting by understanding ZnF-RNA recognition rules for therapeutic development.
Area of Science:
- Molecular Biology
- Biochemistry
- Structural Biology
Background:
- Sequence-specific targeting of endogenous RNAs is crucial for RNA biology and therapeutics.
- RNA-binding zinc fingers (ZnFs) offer potential as designer proteins for RNA targeting due to their modularity.
Purpose of the Study:
- To investigate how RNA-binding ZnF domain sequences dictate binding site specificity.
- To engineer ZnFs with altered RNA sequence preferences for programmable RNA targeting.
Main Methods:
- Systematic mutagenesis of RNA-contacting residues in the ZRANB2 ZnF protein.
- Modified RNA bind-n-seq assay to measure RNA binding affinity and specificity.
- All-atom molecular dynamics simulations to analyze protein-RNA interactions.
Main Results:
- Identified mutant ZnFs exhibiting altered sequence specificity, preferring a GGG motif over the wild-type GGU motif.
- Characterized changes in the protein-RNA hydrogen-bond network correlating with specificity shifts.
- Provided in vitro and in silico insights into ZnF-RNA recognition rules.
Conclusions:
- Understanding ZnF-RNA recognition rules is foundational for developing programmable RNA-targeting tools.
- Engineered ZnFs with modified specificity can be created through targeted mutagenesis.
- This research paves the way for advanced RNA-targeting therapeutics.
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