Zinc-finger recombinase activities in vitro

Prorocic, Marko; Dong, Wenlong; Olorunniji, Femi J.; Akopian, Aram; Schloetel, Jan-Gero; Hannigan, Adele; McPherson, Arlene L.; Stark, W. Marshall

doi:10.1093/nar/gkr652

Cited by 25 publications

(19 citation statements)

References 36 publications

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“…These chimeric enzymes are composed of an activated catalytic domain derived from the resolvase/invertase family of serine recombinases and a zinc-finger DNA-binding domain, which can be custom-designed to recognize almost any DNA sequence (11–16) (Figure 1A). ZFRs catalyse recombination between specific ZFR target sites (17) that consist of two inverted zinc-finger–binding sites (ZFBS) flanking a central 20-bp core sequence recognized by the recombinase catalytic domain (18) (Figure 1B). In contrast to zinc-finger (19–21) and transcription activator-like (TAL) effector nucleases (22,23), ZFRs function autonomously and can excise and integrate transgenes in human and mouse cells without activating the cellular DNA damage response pathway (9,24–26).…”

Section: Introductionmentioning

confidence: 99%

A comprehensive approach to zinc-finger recombinase customization enables genomic targeting in human cells

Gaj

Mercer

Sirk

et al. 2013

Nucleic Acids Research

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Zinc-finger recombinases (ZFRs) represent a potentially powerful class of tools for targeted genetic engineering. These chimeric enzymes are composed of an activated catalytic domain derived from the resolvase/invertase family of serine recombinases and a custom-designed zinc-finger DNA-binding domain. The use of ZFRs, however, has been restricted by sequence requirements imposed by the recombinase catalytic domain. Here, we combine substrate specificity analysis and directed evolution to develop a diverse collection of Gin recombinase catalytic domains capable of recognizing an estimated 3.77 × 107 unique DNA sequences. We show that ZFRs assembled from these engineered catalytic domains recombine user-defined DNA targets with high specificity, and that designed ZFRs integrate DNA into targeted endogenous loci in human cells. This study demonstrates the feasibility of generating customized ZFRs and the potential of ZFR technology for a diverse range of applications, including genome engineering, synthetic biology and gene therapy.

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Section: Introductionmentioning

confidence: 99%

A comprehensive approach to zinc-finger recombinase customization enables genomic targeting in human cells

Gaj

Mercer

Sirk

et al. 2013

Nucleic Acids Research

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“…The inherent disadvantage of using HEases in genome editing is that most of the HEases studied so far tolerate degenerate sequences, and off-target sites with a few base pair mismatches are also cleaved (25,26). ZF recombinases (ZFRs) have also been constructed by fusion of ZF arrays with Tyr or Ser recombinase for ZFR-mediated integration of the donor plasmid in mammalian genomes and site-specific recombination in a bacterial genome (27,28). …”

Section: Introductionmentioning

confidence: 99%

Natural zinc ribbon HNH endonucleases and engineered zinc finger nicking endonuclease

Gupta

2012

Nucleic Acids Research

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Many bacteriophage and prophage genomes encode an HNH endonuclease (HNHE) next to their cohesive end site and terminase genes. The HNH catalytic domain contains the conserved catalytic residues His-Asn-His and a zinc-binding site [CxxC]2. An additional zinc ribbon (ZR) domain with one to two zinc-binding sites ([CxxxxC], [CxxxxH], [CxxxC], [HxxxH], [CxxC] or [CxxH]) is frequently found at the N-terminus or C-terminus of the HNHE or a ZR domain protein (ZRP) located adjacent to the HNHE. We expressed and purified 10 such HNHEs and characterized their cleavage sites. These HNHEs are site-specific and strand-specific nicking endonucleases (NEase or nickase) with 3- to 7-bp specificities. A minimal HNH nicking domain of 76 amino acid residues was identified from Bacillus phage γ HNHE and subsequently fused to a zinc finger protein to generate a chimeric NEase with a new specificity (12–13 bp). The identification of a large pool of previously unknown natural NEases and engineered NEases provides more ‘tools’ for DNA manipulation and molecular diagnostics. The small modular HNH nicking domain can be used to generate rare NEases applicable to targeted genome editing. In addition, the engineered ZF nickase is useful for evaluation of off-target sites in vitro before performing cell-based gene modification.

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“…Enzymes that could be fused to the DBD include epigenetic remodellers that function by modifying DNA or by adding post-translational modifications to histones, such as histone acetyltransferases, HDACs, histone methyltransferases and ubiquitin-conjugating enzymes. Such catalytic domains can be attached to the DBD with flexible or rigid peptide linkers as needed [27,225,226]. …”

Section: Applications In Controlling Gene Networkmentioning

confidence: 99%

Controlling gene networks and cell fate with precision-targeted DNA-binding proteins and small-molecule-based genome readers

Eguchi

Lee

Wan

et al. 2014

Biochemical Journal

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Transcription factors control the fate of a cell by regulating the expression of genes and regulatory networks. Recent successes in inducing pluripotency in terminally differentiated cells as well as directing differentiation with natural transcription factors has lent credence to the efforts that aim to direct cell fate with rationally designed transcription factors. Because DNA-binding factors are modular in design, they can be engineered to target specific genomic sequences and perform pre-programmed regulatory functions upon binding. Such precision-tailored factors can serve as molecular tools to reprogramme or differentiate cells in a targeted manner. Using different types of engineered DNA binders, both regulatory transcriptional controls of gene networks, as well as permanent alteration of genomic content, can be implemented to study cell fate decisions. In the present review, we describe the current state of the art in artificial transcription factor design and the exciting prospect of employing artificial DNA-binding factors to manipulate the transcriptional networks as well as epigenetic landscapes that govern cell fate.

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Zinc-finger recombinase activities in vitro

Cited by 25 publications

References 36 publications

A comprehensive approach to zinc-finger recombinase customization enables genomic targeting in human cells

A comprehensive approach to zinc-finger recombinase customization enables genomic targeting in human cells

Natural zinc ribbon HNH endonucleases and engineered zinc finger nicking endonuclease

Controlling gene networks and cell fate with precision-targeted DNA-binding proteins and small-molecule-based genome readers

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