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Computational design of sequence-specific DNA-binding proteins.

Publication ,  Journal Article
Glasscock, CJ; Pecoraro, R; McHugh, R; Doyle, LA; Chen, W; Boivin, O; Lonnquist, B; Na, E; Politanska, Y; Haddox, HK; Cox, D; Norn, C ...
Published in: bioRxiv
September 21, 2023

Sequence-specific DNA-binding proteins (DBPs) play critical roles in biology and biotechnology, and there has been considerable interest in the engineering of DBPs with new or altered specificities for genome editing and other applications. While there has been some success in reprogramming naturally occurring DBPs using selection methods, the computational design of new DBPs that recognize arbitrary target sites remains an outstanding challenge. We describe a computational method for the design of small DBPs that recognize specific target sequences through interactions with bases in the major groove, and employ this method in conjunction with experimental screening to generate binders for 5 distinct DNA targets. These binders exhibit specificity closely matching the computational models for the target DNA sequences at as many as 6 base positions and affinities as low as 30-100 nM. The crystal structure of a designed DBP-target site complex is in close agreement with the design model, highlighting the accuracy of the design method. The designed DBPs function in both Escherichia coli and mammalian cells to repress and activate transcription of neighboring genes. Our method is a substantial step towards a general route to small and hence readily deliverable sequence-specific DBPs for gene regulation and editing.

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bioRxiv

DOI

Publication Date

September 21, 2023

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United States
 

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Glasscock, C. J., Pecoraro, R., McHugh, R., Doyle, L. A., Chen, W., Boivin, O., … Baker, D. (2023). Computational design of sequence-specific DNA-binding proteins. BioRxiv. https://doi.org/10.1101/2023.09.20.558720
Glasscock, Cameron J., Robert Pecoraro, Ryan McHugh, Lindsey A. Doyle, Wei Chen, Olivier Boivin, Beau Lonnquist, et al. “Computational design of sequence-specific DNA-binding proteins.BioRxiv, September 21, 2023. https://doi.org/10.1101/2023.09.20.558720.
Glasscock CJ, Pecoraro R, McHugh R, Doyle LA, Chen W, Boivin O, et al. Computational design of sequence-specific DNA-binding proteins. bioRxiv. 2023 Sep 21;
Glasscock, Cameron J., et al. “Computational design of sequence-specific DNA-binding proteins.BioRxiv, Sept. 2023. Pubmed, doi:10.1101/2023.09.20.558720.
Glasscock CJ, Pecoraro R, McHugh R, Doyle LA, Chen W, Boivin O, Lonnquist B, Na E, Politanska Y, Haddox HK, Cox D, Norn C, Coventry B, Goreshnik I, Vafeados D, Lee GR, Gordan R, Stoddard BL, DiMaio F, Baker D. Computational design of sequence-specific DNA-binding proteins. bioRxiv. 2023 Sep 21;

Published In

bioRxiv

DOI

Publication Date

September 21, 2023

Location

United States