ArticleProceedings of the National Academy of Sciences of the United States of America2015
Balancing between affinity and speed in target DNA search by zinc-finger proteins via modulation of dynamic conformational ensemble.
Article in Proceedings of the National Academy of Sciences of the United States of America, 2015. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 67 papers.
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Who cites it
67 citing papers in PubMed, 111 citations in OpenAlex.
- Observation of hidden dynamics underlying facilitated dissociation of proteins from DNA.Chemical communications (Cambridge, England) · 2026Article
- Bipartite DNA binding domain of transcription factor BCL11B binds clustered short DNA sequence motifs.bioRxiv : the preprint server for biology · 2026Article
- Mechanisms for DNA Interplay in Eukaryotic Transcription Factors.Annual review of biophysics · 2025Review
- Protein and DNA Conformational Changes Contribute to Specificity of Cre Recombinase.Biochemistry · 2025Article
- Protein and DNA Conformational Changes Contribute to Specificity of Cre Recombinase.bioRxiv : the preprint server for biology · 2024Article
- Dynamics in Cre-loxP site-specific recombination.Current opinion in structural biology · 2024Review
- Structure and RNA-binding of the helically extended Roquin CCCH-type zinc finger.Nucleic acids research · 2024Article
- Article
- Skipping events impose repeated binding attempts: profound kinetic implications of protein-DNA conformational changes.Nucleic acids research · 2024Article
- Ordered and disordered regions of the Origin Recognition Complex direct differential in vivo binding at distinct motif sequences.Nucleic acids research · 2024Article
- Analyzing paramagnetic NMR data on target DNA search by proteins using a discrete-state kinetic model for translocation.Biopolymers · 2024Article
- smFRET Detection of Cis and Trans DNA Interactions by the BfiI Restriction Endonuclease.The journal of physical chemistry. B · 2023Article
- Negatively charged, intrinsically disordered regions can accelerate target search by DNA-binding proteins.Nucleic acids research · 2023Article
- Conformational Change of Transcription Factors from Search to Specific Binding: AThe journal of physical chemistry. B · 2022Article
- Flexible Target Recognition of the Intrinsically Disordered DNA-Binding Domain of CytR Monitored by Single-Molecule Fluorescence Spectroscopy.The journal of physical chemistry. B · 2022Article
- Cognate DNA Recognition by Engrailed Homeodomain Involves a Conformational Change Controlled via an Electrostatic-Spring-Loaded Latch.International journal of molecular sciences · 2022Article
- Kinetic origin of nucleosome invasion by pioneer transcription factors.Biophysical journal · 2021Article
- Single molecule characterization of the binding kinetics of a transcription factor and its modulation by DNA sequence and methylation.Nucleic acids research · 2021Article
- Engineering of the genome editing protein Cas9 to slide along DNA.Scientific reports · 2021Article
- What Are the Molecular Requirements for Protein Sliding along DNA?The journal of physical chemistry. B · 2021Article
7 more citing papers are in PubMed but not listed here.
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Authors and funding
6 authors at 2 institutions in 2 countries.
Funding
Abstract
Although engineering of transcription factors and DNA-modifying enzymes has drawn substantial attention for artificial gene regulation and genome editing, most efforts focus on affinity and specificity of the DNA-binding proteins, typically overlooking the kinetic properties of these proteins. However, a simplistic pursuit of high affinity can lead to kinetically deficient proteins that spend too much time at nonspecific sites before reaching their targets on DNA. We demonstrate that structural dynamic knowledge of the DNA-scanning process allows for kinetically and thermodynamically balanced engineering of DNA-binding proteins. Our current study of the zinc-finger protein Egr-1 (also known as Zif268) and its nuclease derivatives reveals kinetic and thermodynamic roles of the dynamic conformational equilibrium between two modes during the DNA-scanning process: one mode suitable for search and the other for recognition. By mutagenesis, we were able to shift this equilibrium, as confirmed by NMR spectroscopy. Using fluorescence and biochemical assays as well as computational simulations, we analyzed how the shifts of the conformational equilibrium influence binding affinity, target search kinetics, and efficiency in displacing other proteins from the target sites. A shift toward the recognition mode caused an increase in affinity for DNA and a decrease in search efficiency. In contrast, a shift toward the search mode caused a decrease in affinity and an increase in search efficiency. This accelerated site-specific DNA cleavage by the zinc-finger nuclease, without enhancing off-target cleavage. Our study shows that appropriate modulation of the dynamic conformational ensemble can greatly improve zinc-finger technology, which has used Egr-1 (Zif268) as a major scaffold for engineering.
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