Structure and function of novel prokaryotic DNA transposases

National Institute Of Diabetes And Digestive And Kidney Diseases

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Abstract

We have been investigating Insertion Sequence (IS) movements in multidrug resistant bacteria with a focus on carbapenemase-producing Enterobacteriaceae (CPE). We have previously analyzed the genomic contexts of ISs in several clinical and surveillance CPE isolates from the NIH Clinical Center, using target site duplications (TSDs) and their distribution patterns as guides, and discovered that a large fraction of plasmid reorganizations result from IS26 intramolecular replicative transpositions, including replicon fusions, DNA inversions, and deletions (1,2). We are interested in learning about the mechanisms behind these DNA rearrangements, and are currently investigating the biochemical and biophysical properties of a variety of recombinantly expressed replicative transposases. Our recent focus has been on a transposase from the IS256 family that is mobilized by the so-called "copy-out paste-in" mechanism (1,2), the ISCth4 transposase whose structure we have solved using X-ray crystallography in complex with various DNA substrates (3). The results show that it forms an unusual asymmetric dimeric transpososome in which an array of N-terminal domains binds a single transposon end whilst the catalytic domain moves to accommodate its varying substrates. This asymmetric arrangement explains the unusual asymmetric cleavage of the transposon ends that is a hallmark of copy-out-paste-in transposition. 1. He, Hickman, Varani, Siguier, Chandler, Dekker, and Dyda (2015) Insertion Sequence IS26 reorganizes plasmids in clinically isolated multidrug-resistant bacteria by replicative transposition. mBio 3, e00762-15. 2. He, Chandler, Varani, Hickman, Dekker, and Dyda (2016) Mechanism of evolution in high-consequence drug resistance plasmids. mBio 6, e01987. 3. Kosek D, Hickman AB, Ghirlando R, He S, Dyda F (2021). Structures of ISCth4 transpososomes reveal the role of asymmetry in copy-out/paste-in DNA transposition. EMBO J 40, e105666. https://doi.org/10.15252/embj.2020105666