title: Conformational toggling controls target site choice for the heteromeric transposase element Tn7.
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.
authors.affiliation: Department of Microbiology, Cornell University, Ithaca, NY 14853, USA.
authors.forename: Qiaojuan
authors.initials: Q
authors.lastname: Shi
authors.affiliation: Department of Microbiology, Cornell University, Ithaca, NY 14853, USA.
authors.forename: Marco R
authors.initials: MR
authors.lastname: Straus
authors.affiliation: Department of Biochemistry and Biomedical Sciences, McMaster University, Hamilton, Ontario, L8S 4K1, Canada.
authors.forename: Jeremy J
authors.initials: JJ
authors.lastname: Caron
authors.affiliation: Department of Biochemistry and Biomedical Sciences, McMaster University, Hamilton, Ontario, L8S 4K1, Canada.
authors.forename: Huasheng
authors.initials: H
authors.lastname: Wang
authors.affiliation: Department of Biochemistry and Biomedical Sciences, McMaster University, Hamilton, Ontario, L8S 4K1, Canada.
authors.forename: Yu Seon
authors.initials: YS
authors.lastname: Chung
authors.affiliation: Department of Biochemistry and Biomedical Sciences, McMaster University, Hamilton, Ontario, L8S 4K1, Canada guarnea@mcmaster.ca.
authors.forename: Alba
authors.initials: A
authors.lastname: Guarné
authors.affiliation: Department of Microbiology, Cornell University, Ithaca, NY 14853, USA jep48@cornell.edu.
authors.forename: Joseph E
authors.initials: JE
authors.lastname: Peters
mesh_terms: D000409:Alanine / Q000737:chemistry; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D004247:DNA / Q000378:metabolism; D004251:DNA Transposable Elements; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D008958:Models, Molecular; D009154:Mutation; D017510:Protein Folding; D017434:Protein Structure, Tertiary; D019895:Transposases / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D004251:DNA Transposable Elements; D004268:DNA-Binding Proteins; D004247:DNA; D019895:Transposases; D000409:Alanine