Engineered Transcription Activator-Like Effector Dimer Proteins Confer DNA Loop-Dependent Gene Repression Comparable to Lac Repressor

Authors

Nicole A. Becker
Justin P. Peters, University of Northern IowaFollow
Elizabeth Lewis, Mayo Clinic Graduate School of Biomedical Sciences
Camden L. Daby, Mayo Clinic Graduate School of Biomedical Sciences
Karl Clark, Mayo Clinic Graduate School of Biomedical Sciences
L. James Maher, Mayo Clinic Graduate School of Biomedical SciencesFollow

Comments

First published in Nucleic Acids Research, v52i16 (Sept 2024) published by Oxford University Press. DOI: https://doi.org/10.1093/nar/gkae656

Document Type

Article

Publication Version

Published Version

Journal/Book/Conference Title

Nucleic Acids Research

Volume

52

Issue

16

First Page

9996

Last Page

10004

Abstract

Natural prokaryotic gene repression systems often exploit DNA looping to increase the local concentration of gene repressor proteins at a regulated promoter via contributions from repressor proteins bound at distant sites. Using principles from the Escherichia coli lac operon we design analogous repression systems based on target sequence-programmable Transcription Activator-Like Effector dimer (TALED) proteins. Such engineered switches may be valuable for synthetic biology and therapeutic applications. Previous TALEDs with inducible non-covalent dimerization showed detectable, but limited, DNA loop-based repression due to the repressor protein dimerization equilibrium. Here, we show robust DNA loop-dependent bacterial promoter repression by covalent TALEDs and verify that DNA looping dramatically enhances promoter repression in E. coli. We characterize repression using a thermodynamic model that quantitates this favorable contribution of DNA looping. This analysis unequivocally and quantitatively demonstrates that optimized TALED proteins can drive loop-dependent promoter repression in E. coli comparable to the natural LacI repressor system. This work elucidates key design principles that set the stage for wide application of TALED-dependent DNA loop-based repression of target genes.

Department

Department of Chemistry and Biochemistry

Original Publication Date

9-9-2024

Object Description

1 PDF File

DOI of published version

10.1093/nar/gkae656

Repository

UNI ScholarWorks, Rod Library, University of Northern Iowa

Copyright

©2024 The Author(s) This is an Open Access article distributed under the terms of the Creative Commons Attribution-NonCommercial License (https://creativecommons.org/licenses/by-nc/4.0/).

Creative Commons License

This work is licensed under a Creative Commons Attribution-NonCommercial 4.0 International License

Language

en

File Format

application/pdf

Recommended Citation

Becker, Nicole A.; Peters, Justin P.; Lewis, Elizabeth; Daby, Camden L.; Clark, Karl; and Maher, L. James, "Engineered Transcription Activator-Like Effector Dimer Proteins Confer DNA Loop-Dependent Gene Repression Comparable to Lac Repressor" (2024). Faculty Publications. 6502.
https://scholarworks.uni.edu/facpub/6502