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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
PubMed
Summary
This summary is machine-generated.

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.

Keywords:
RBNSRNA-binding zinc fingerZRANB2rational design

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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.