Unlocking Tn3-family transposase activity in vitro unveils an asymetric pathway for transposome assembly

Emilien Nicolas, Cédric A Oger, Nathan Nguyen, Michaël Lambin, Amandine Draime, Sébastien C Leterme, Michael Chandler, Bernard F J Hallet

Research output: Contribution to journalArticlepeer-review

Abstract

The Tn3 family is a widespread group of replicative transposons that are notorious for their contribution to the dissemination of antibiotic resistance and the emergence of multiresistant pathogens worldwide. The TnpA transposase of these elements catalyzes DNA breakage and rejoining reactions required for transposition. It also is responsible for target immunity, a phenomenon that prevents multiple insertions of the transposon into the same genomic region. However, the molecular mechanisms whereby TnpA acts in both processes remain unknown. Here, we have developed sensitive biochemical assays for the TnpA transposase of the Tn3-family transposon Tn4430 and used these assays to characterize previously isolated TnpA mutants that are selectively affected in immunity. Compared with wild-type TnpA, these mutants exhibit deregulated activities. They spontaneously assemble a unique asymmetric synaptic complex in which one TnpA molecule simultaneously binds two transposon ends. In this complex, TnpA is in an activated state competent for DNA cleavage and strand transfer. Wild-type TnpA can form this complex only on precleaved ends mimicking the initial step of transposition. The data suggest that transposition is controlled at an early stage of transpososome assembly, before DNA cleavage, and that mutations affecting immunity have unlocked TnpA by stabilizing the protein in a monomeric activated synaptic configuration. We propose an asymmetric pathway for coupling active transpososome assembly with proper target recruitment and discuss this model with respect to possible immunity mechanisms.

Original languageEnglish
Pages (from-to)E669-E678
JournalProceedings of the National Academy of Sciences of the United States of America
Volume114
Issue number5
DOIs
Publication statusPublished - 31 Jan 2017
Externally publishedYes

Keywords

  • DNA/chemistry
  • DNA Transposable Elements
  • Escherichia coli/genetics
  • Mutation
  • Transposases/chemistry

Fingerprint Dive into the research topics of 'Unlocking Tn3-family transposase activity in vitro unveils an asymetric pathway for transposome assembly'. Together they form a unique fingerprint.

Cite this