Development of Gene-Targeted Polypyridyl Triplex-Forming Oligonucleotide Hybrids.
Chembiochem
; 21(24): 3563-3574, 2020 12 11.
Article
em En
| MEDLINE
| ID: mdl-32755000
ABSTRACT
In the field of nucleic acid therapy there is major interest in the development of libraries of DNA-reactive small molecules which are tethered to vectors that recognize and bind specific genes. This approach mimics enzymatic gene editors, such as ZFNs, TALENs and CRISPR-Cas, but overcomes the limitations imposed by the delivery of a large protein endonuclease which is required for DNA cleavage. Here, we introduce a chemistry-based DNA-cleavage system comprising an artificial metallo-nuclease (AMN) that oxidatively cuts DNA, and a triplex-forming oligonucleotide (TFO) that sequence-specifically recognises duplex DNA. The AMN-TFO hybrids coordinate CuII ions to form chimeric catalytic complexes that are programmable - based on the TFO sequence employed - to bind and cut specific DNA sequences. Use of the alkyne-azide cycloaddition click reaction allows scalable and high-throughput generation of hybrid libraries that can be tuned for specific reactivity and gene-of-interest knockout. As a first approach, we demonstrate targeted cleavage of purine-rich sequences, optimisation of the hybrid system to enhance stability, and discrimination between target and off-target sequences. Our results highlight the potential of this approach where the cutting unit, which mimics the endonuclease cleavage machinery, is directly bound to a TFO guide by click chemistry.
Palavras-chave
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Oligonucleotídeos
/
DNA
/
Cobre
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Endonucleases
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Metaloproteínas
Idioma:
En
Revista:
Chembiochem
Assunto da revista:
BIOQUIMICA
Ano de publicação:
2020
Tipo de documento:
Article
País de afiliação:
Irlanda