en
Scientific article
Open access
English

Targeting IS608 transposon integration to highly specific sequences by structure-based transposon engineering

Published inNucleic acids research, vol. 46, no. 8, p. 4152-4163
Publication date2018-05-04
First online date2018-04-09
Abstract

Transposable elements are efficient DNA carriers and thus important tools for transgenesis and insertional mutagenesis. However, their poor target sequence specificity constitutes an important limitation for site-directed applications. The insertion sequence IS 608 from Helicobacter pylori recognizes a specific tetranucleotide sequence by base pairing, and its target choice can be re-programmed by changes in the transposon DNA. Here, we present the crystal structure of the IS608 target capture complex in an active conformation, providing a complete picture of the molecular interactions between transposon and target DNA prior to integration. Based on this, we engineered IS608 variants to direct their integration specifically to various 12/17-nt long target sites by extending the base pair interaction net- work between the transposon and the target DNA. We demonstrate in vitro that the engineered transposons efficiently select their intended target sites. Our data further elucidate how the distinct secondary structure of the single-stranded transposon intermediate prevents extended target specificity in the wild-type transposon, allowing it to move between diverse genomic sites. Our strategy enables efficient targeting of unique DNA sequences with high specificity in an easily programmable manner, opening possibilities for the use of the IS608 system for site-specific gene insertions.

eng
Affiliation Not a UNIGE publication
Funding
  • European Commission - European Molecular Biology Laboratory Interdisciplinary Postdocs [229597]
Citation (ISO format)
MORERO, Natalia Rosalía et al. Targeting IS<i>608</i> transposon integration to highly specific sequences by structure-based transposon engineering. In: Nucleic acids research, 2018, vol. 46, n° 8, p. 4152–4163. doi: 10.1093/nar/gky235
Main files (1)
Article (Published version)
Identifiers
ISSN of the journal0305-1048
12views
1downloads

Technical informations

Creation03/12/2024 5:10:03 PM
First validation03/28/2024 3:00:52 PM
Update time03/28/2024 3:00:52 PM
Status update03/28/2024 3:00:52 PM
Last indexation03/28/2024 3:01:12 PM
All rights reserved by Archive ouverte UNIGE and the University of GenevaunigeBlack