Structural heterogeneity of attC integron recombination sites revealed by optical tweezers

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Contributors

Abstract

A predominant tool for adaptation in Gram-negative bacteria is the functional genetic platform called integron. Integrons capture and rearrange promoterless gene cassettes in a unique recombination process involving the recognition of folded single-stranded DNA hairpinsso-called attC siteswith a strong preference for the attC bottom strand. While structural elements have been identified to promote this preference, their mechanistic action remains incomplete. Here, we used high-resolution single-molecule optical tweezers (OT) to characterize secondary structures formed by the attC bottom () and top () strands of the paradigmatic attC(aadA7) site. We found for both sequences two structuresa straight, canonical hairpin and a kinked hairpin. Remarkably, the recombination-preferred predominantly formed the straight hairpin, while the preferentially adopted the kinked structure, which exposes only one complete recombinase binding box. By a mutational analysis, we identified three bases in the unpaired central spacer, which could invert the preferred conformations and increase the recombination frequency of the in vivo. A bioinformatics screen revealed structural bias toward a straight, canonical hairpin conformation in the bottom strand of many antibiotic resistance cassettes attC sites. Thus, we anticipate that structural fine tuning could be a mechanism in many biologically active DNA hairpins.

Details

Original languageEnglish
Pages (from-to)1861-1870
Number of pages10
JournalNucleic acids research
Volume47
Issue number4
Publication statusPublished - 28 Feb 2019
Peer-reviewedYes

External IDs

Scopus 85062287208

Keywords

Keywords

  • SINGLE-STRANDED-DNA, CASSETTE INSERTION, ELASTIC PROPERTIES, MOLECULES, BINDS