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A single active site in the mariner transposase cleaves DNA strands of opposite polarity

Claeys Bouuaert, Corentin; Chalmers, Ronald

Authors

Corentin Claeys Bouuaert claeysbc@mskcc.org

RONALD CHALMERS RONALD.CHALMERS@NOTTINGHAM.AC.UK
Professor of Biochemistry and Cell Biology



Abstract

The RNase H structural fold defines a large family of nucleic acid metabolizing enzymes that catalyze phosphoryl transfer reactions using two divalent metal ions in the active site. Almost all of these reactions involve only one strand of the nucleic acid substrates. In contrast, cut-and-paste transposases cleave two DNA strands of opposite polarity, which is usually achieved via an elegant hairpin mechanism. In the mariner transposons, the hairpin intermediate is absent and key aspects of the mechanism by which the transposon ends are cleaved remained unknown. Here, we characterize complexes involved prior to catalysis, which define an asymmetric pathway for transpososome assembly. Using mixtures of wild-type and catalytically inactive transposases, we show that all the catalytic steps of transposition occur within the context of a dimeric transpososome. Crucially, we find that each active site of a transposase dimer is responsible for two hydrolysis and one transesterification reaction at the same transposon end. These results provide the first strong evidence that a DDE/D active site can hydrolyze DNA strands of opposite polarity, a mechanism that has rarely been observed with any type of nuclease.

Citation

Claeys Bouuaert, C., & Chalmers, R. (in press). A single active site in the mariner transposase cleaves DNA strands of opposite polarity. Nucleic Acids Research, gkx826, https://doi.org/10.1093/nar/gkx826

Journal Article Type Article
Acceptance Date Sep 8, 2017
Online Publication Date Sep 19, 2017
Deposit Date Nov 6, 2017
Publicly Available Date Nov 6, 2017
Journal Nucleic Acids Research
Print ISSN 0305-1048
Electronic ISSN 1362-4962
Publisher Oxford University Press
Peer Reviewed Peer Reviewed
Volume gkx826
DOI https://doi.org/10.1093/nar/gkx826
Public URL http://eprints.nottingham.ac.uk/id/eprint/47838
Publisher URL https://doi.org/10.1093/nar/gkx826
Copyright Statement Copyright information regarding this work can be found at the following address: http://creativecommons.org/licenses/by/4.0

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Copyright Statement
Copyright information regarding this work can be found at the following address: http://creativecommons.org/licenses/by/4.0





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