ArticleNucleic acids research1997
Purification, biochemical characterization and protein-DNA interactions of the I-CreI endonuclease produced in Escherichia coli.
Article in Nucleic acids research, 1997. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 21 papers.
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Who cites it
21 citing papers in PubMed, 46 citations in OpenAlex.
- F-CphI represents a new homing endonuclease family using the Endo VII catalytic motif.Mobile DNA · 2018Article
- Targeted mutagenesis of a conserved anther-expressed P450 gene confers male sterility in monocots.Plant biotechnology journal · 2017Article
- Crystal Structure of the Homing Endonuclease I-CvuI Provides a New Template for Genome Modification.The Journal of biological chemistry · 2015Article
- 5'-Cytosine-phosphoguanine (CpG) methylation impacts the activity of natural and engineered meganucleases.The Journal of biological chemistry · 2012Article
- Non-specific protein-DNA interactions control I-CreI target binding and cleavage.Nucleic acids research · 2012Article
- Double-strand breaks repair by gene conversion in silkworm holocentric chromosomes.Molecular genetics and genomics : MGG · 2011Article
- Molecular basis of engineered meganuclease targeting of the endogenous human RAG1 locus.Nucleic acids research · 2011Article
- Homing endonucleases: from basics to therapeutic applications.Cellular and molecular life sciences : CMLS · 2010Review
- Mutational analysis of active-site residues in the Mycobacterium leprae RecA intein, a LAGLIDADG homing endonuclease: Asp(122) and Asp(193) are crucial to the double-stranded DNA cleavage activity whereas Asp(218) is not.Protein science : a publication of the Protein Society · 2010Article
- Characterization of Mycobacterium leprae RecA intein, a LAGLIDADG homing endonuclease, reveals a unique mode of DNA binding, helical distortion, and cleavage compared with a canonical LAGLIDADG homing endonuclease.The Journal of biological chemistry · 2009Article
- Biochemical and mutagenic analysis of I-CreII reveals distinct but important roles for both the H-N-H and GIY-YIG motifs.Nucleic acids research · 2009Article
- Crystal structures of I-SceI complexed to nicked DNA substrates: snapshots of intermediates along the DNA cleavage reaction pathway.Nucleic acids research · 2008Article
- Computer design of obligate heterodimer meganucleases allows efficient cutting of custom DNA sequences.Nucleic acids research · 2008Article
- Crystallization and preliminary X-ray diffraction analysis on the homing endonuclease I-Dmo-I in complex with its target DNA.Acta crystallographica. Section F, Structural biology and crystallization communications · 2007Article
- The C-terminal loop of the homing endonuclease I-CreI is essential for site recognition, DNA binding and cleavage.Nucleic acids research · 2007Article
- Mycobacterium tuberculosis RecA intein, a LAGLIDADG homing endonuclease, displays Mn(2+) and DNA-dependent ATPase activity.Nucleic acids research · 2003Article
- A novel engineered meganuclease induces homologous recombination in yeast and mammalian cells.Nucleic acids research · 2003Article
- Mutations altering the cleavage specificity of a homing endonuclease.Nucleic acids research · 2002Article
- Rapid evolution of the DNA-binding site in LAGLIDADG homing endonucleases.Nucleic acids research · 2001Article
- Biochemical characterization of I-CmoeI reveals that this H-N-H homing endonuclease shares functional similarities with H-N-H colicins.Nucleic acids research · 2000Article
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Authors and funding
4 authors at 1 institution in 1 country.
Funding
No grant is acknowledged in the PubMed record.
Abstract
I- CreI is a member of the LAGLI-DADG family of homing nucleases; however, unlike most members of this family it contains only a single copy of this signature motif. I- CreI was over-expressed in Escherichia coli, and a simple purification protocol developed that gave reasonably pure protein in high yield. Size-exclusion chromatography and chemical cross-linking indicated that the protein is a dimer in solution. DNA cleavage by I- CreI was absolutely dependent on Mg2+(or Mn2+), and was inhibited by monovalent cations. I- CreI displayed a surprisingly high temperature optimum (>50 degrees C), with full activity occurring even at 70 degrees C. Interestingly, SDS was needed for efficient release of the cleavage products from the protein, indicating formation of very stable DNA-protein complexes. In contrast to these robust characteristics, purified I- CreI was unstable; however, it could be stabilized by the addition of either target or non-target DNA. Mobility shift assays revealed that I- CreI binds to DNA in the absence of Mg2+. Hydroxyl radical footprinting showed that I- CreI strongly protected the backbone of a continuous stretch of at least 12 nt on each strand that were shifted, relative to each other, by 2 bp in the 3'direction. Methylation protection and interference analyses were also performed, and together with the hydroxyl radical footprinting, indicate that I- CreI binds in both the major and minor grooves of its target DNA.
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