ArticleNucleic acids research2012
Creating highly specific nucleases by fusion of active restriction endonucleases and catalytically inactive homing endonucleases.
Article in Nucleic acids research, 2012. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 13 papers.
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Who cites it
13 citing papers in PubMed.
- Mapping the zygote-to-adult developmental cell phylogeny in Arabidopsis thaliana reveals a three-cell rule of branching.Nature plants · 2026Article
- Mapping single-cell-resolution cell phylogeny reveals cell population dynamics during organ development.Nature methods · 2021Article
- Applications of Alternative Nucleases in the Age of CRISPR/Cas9.International journal of molecular sciences · 2017Review
- DNA cleavage enzymes for treatment of persistent viral infections: recent advances and the pathway forward.Virology · 2014Review
- Homing endonucleases: from genetic anomalies to programmable genomic clippers.Methods in molecular biology (Clifton, N.J.) · 2014Review
- Long distance multiple-site directed plasmid mutagenesis by one-step PCR using non-overlapped primers.Molecular biotechnology · 2013Article
- The monomeric GIY-YIG homing endonuclease I-BmoI uses a molecular anchor and a flexible tether to sequentially nick DNA.Nucleic acids research · 2013Article
- Site- and strand-specific nicking of DNA by fusion proteins derived from MutH and I-SceI or TALE repeats.Nucleic acids research · 2013Article
- TALE-PvuII fusion proteins--novel tools for gene targeting.PloS one · 2013Article
- Compact designer TALENs for efficient genome engineering.Nature communications · 2013Article
- EENdb: a database and knowledge base of ZFNs and TALENs for endonuclease engineering.Nucleic acids research · 2013Article
- Monomeric site-specific nucleases for genome editing.Proceedings of the National Academy of Sciences of the United States of America · 2012Article
- A novel zinc-finger nuclease platform with a sequence-specific cleavage module.Nucleic acids research · 2012Article
Corrections and comments
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Authors and funding
4 authors.
Funding
No grant is acknowledged in the PubMed record.
Abstract
Zinc-finger nucleases and TALE nucleases are produced by combining a specific DNA-binding module and a non-specific DNA-cleavage module, resulting in nucleases able to cleave DNA at a unique sequence. Here a new approach for creating highly specific nucleases was pursued by fusing a catalytically inactive variant of the homing endonuclease I-SceI, as DNA binding-module, to the type IIP restriction enzyme PvuII, as cleavage module. The fusion enzymes were designed to recognize a composite site comprising the recognition site of PvuII flanked by the recognition site of I-SceI. In order to reduce activity on PvuII sites lacking the flanking I-SceI sites, the enzymes were optimized so that the binding of I-SceI to its sites positions PvuII for cleavage of the composite site. This was achieved by optimization of the linker and by introducing amino acid substitutions in PvuII which decrease its activity or disturb its dimer interface. The most specific variant showed a more than 1000-fold preference for the addressed composite site over an unaddressed PvuII site. These results indicate that using a specific restriction enzyme, such as PvuII, as cleavage module, offers an alternative to the otherwise often used catalytic domain of FokI, which by itself does not contribute to the specificity of the engineered nuclease.
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