Formation of a nucleoprotein complex containing Tn7 and its target DNA regulates transposition initiation

Zachary Skelding, Robert Sarnovsky, Nancy L. Craig

Research output: Contribution to journalArticle

Abstract

Tn7 insertion into its specific target site, attTn7, is mediated by the proteins TnsA, TnsB, TnsC and TnsD. The double-strand breaks that separate Tn7 from the donor DNA require the Tns proteins, the transposon and an attTn7 target DNA, suggesting that a prerequisite for transposition is the formation of a nucleoprotein complex containing TnsABC+D, and these DNAs. Here, we identify a TnsABC+D transposon-attTn7 complex, and demonstrate that it is a transposition intermediate. We demonstrate that an interaction between TnsB, the transposase subunit that binds to the transposon ends, and TnsC, the target DNA-binding protein that controls the activity of the transposase, is essential for assembly of the TnsABC+D transposon-attTn7 complex. We also show that certain TnsB residues are required for recombination because they mediate a TnsB-TnsC interaction critical to formation of the TnsABC+D transposon-attTn7 complex. We demonstrate that TnsA, the other transposase subunit, which also interacts with TnsC, greatly stabilizes the TnsABC+D transposon-attTn7 complex. Thus multiple interactions between the transposase subunits, TnsA and TnsB, and the target-binding transposase activator, TnsC, control Tn7 transposition.

Original languageEnglish (US)
Pages (from-to)3494-3504
Number of pages11
JournalEMBO Journal
Volume21
Issue number13
DOIs
StatePublished - Jul 1 2002

Keywords

  • DNA transposition
  • DNA-protein complex
  • Enzyme activation
  • Tn7
  • Transposase

ASJC Scopus subject areas

  • Neuroscience(all)
  • Molecular Biology
  • Biochemistry, Genetics and Molecular Biology(all)
  • Immunology and Microbiology(all)

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