Expanding the binding specificity for RNA recognition by a PUF domain

Wei Zhou, Daniel Melamed, Gabor Banyai, Cindy Meyer, Thomas Tuschl, Marvin Wickens, Junyue Cao () and Stanley Fields ()
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Wei Zhou: University of Washington, Seattle
Daniel Melamed: University of Washington, Seattle
Gabor Banyai: The Rockefeller University
Cindy Meyer: The Rockefeller University
Thomas Tuschl: The Rockefeller University
Marvin Wickens: University of Wisconsin-Madison
Junyue Cao: The Rockefeller University
Stanley Fields: University of Washington, Seattle

Nature Communications, 2021, vol. 12, issue 1, 1-15

Abstract: Abstract The ability to design a protein to bind specifically to a target RNA enables numerous applications, with the modular architecture of the PUF domain lending itself to new RNA-binding specificities. For each repeat of the Pumilio-1 PUF domain, we generate a library that contains the 8,000 possible combinations of amino acid substitutions at residues critical for RNA contact. We carry out yeast three-hybrid selections with each library against the RNA recognition sequence for Pumilio-1, with any possible base present at the position recognized by the randomized repeat. We use sequencing to score the binding of each variant, identifying many variants with highly repeat-specific interactions. From these data, we generate an RNA binding code specific to each repeat and base. We use this code to design PUF domains against 16 RNAs, and find that some of these domains recognize RNAs with two, three or four changes from the wild type sequence.

Date: 2021
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Persistent link: https://EconPapers.repec.org/RePEc:nat:natcom:v:12:y:2021:i:1:d:10.1038_s41467-021-25433-6

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DOI: 10.1038/s41467-021-25433-6

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