Chemical Approach to Biological Safety: Molecular-Level Control of an Integrated Zinc Finger Nuclease

Eszter Németh, Masamitsu N Asaka, Kohsuke Kato, Zita Fábián, Chris Oostenbrink, Hans Erik Mølager Christensen, Kyosuke Nagata, Béla Gyurcsik*

*Corresponding author for this work

Research output: Contribution to journalJournal articleResearchpeer-review


Application of artificial nucleases (ANs) in genome editing is still hindered by their cytotoxicity related to off-target cleavages. This problem can be targeted by regulation of the nuclease domain. Here, we provide an experimental survey of computationally designed integrated zinc finger nucleases, constructed by linking the inactivated catalytic centre and the allosteric activator sequence of the colicin E7 nuclease domain to the two opposite termini of a zinc finger array. DNA specificity and metal binding were confirmed by electrophoretic mobility shift assays, synchrotron radiation circular dichroism spectroscopy, and nano-electrospray ionisation mass spectrometry. In situ intramolecular activation of the nuclease domain was observed, resulting in specific cleavage of DNA with moderate activity. This study represents a new approach to AN design through integrated nucleases consisting of three (regulator, DNA-binding, and nuclease) units, rather than simple chimera. The optimisation of such ANs could lead to safe gene editing enzymes.
Original languageEnglish
Issue number1
Pages (from-to)66-75
Publication statusPublished - 2018


Dive into the research topics of 'Chemical Approach to Biological Safety: Molecular-Level Control of an Integrated Zinc Finger Nuclease'. Together they form a unique fingerprint.

Cite this