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Nucleic Acids Res. 2015 Dec 15;43(22):10734-45. doi: 10.1093/nar/gkv913. Epub 2015 Sep 17.

Conformational toggling controls target site choice for the heteromeric transposase element Tn7.

Author information

1
Department of Microbiology, Cornell University, Ithaca, NY 14853, USA.
2
Department of Biochemistry and Biomedical Sciences, McMaster University, Hamilton, Ontario, L8S 4K1, Canada.
3
Department of Biochemistry and Biomedical Sciences, McMaster University, Hamilton, Ontario, L8S 4K1, Canada guarnea@mcmaster.ca.
4
Department of Microbiology, Cornell University, Ithaca, NY 14853, USA jep48@cornell.edu.

Abstract

The bacterial transposon Tn7 facilitates horizontal transfer by directing transposition into actively replicating DNA with the element-encoded protein TnsE. Structural analysis of the C-terminal domain of wild-type TnsE identified a novel protein fold including a central V-shaped loop that toggles between two distinct conformations. The structure of a robust TnsE gain-of-activity variant has this loop locked in a single conformation, suggesting that conformational flexibility regulates TnsE activity. Structure-based analysis of a series of TnsE mutants relates transposition activity to DNA binding stability. Wild-type TnsE appears to naturally form an unstable complex with a target DNA, whereas mutant combinations required for large changes in transposition frequency and targeting stabilized this interaction. Collectively, our work unveils a unique structural proofreading mechanism where toggling between two conformations regulates target commitment by limiting the stability of target DNA engagement until an appropriate insertion site is identified.

PMID:
26384427
PMCID:
PMC4678854
DOI:
10.1093/nar/gkv913
[Indexed for MEDLINE]
Free PMC Article

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