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The Mu transpososome structure sheds light on DDE recombinase evolution

Abstract

Studies of bacteriophage Mu transposition paved the way for understanding retroviral integration and V(D)J recombination as well as many other DNA transposition reactions. Here we report the structure of the Mu transpososome—Mu transposase (MuA) in complex with bacteriophage DNA ends and target DNA—determined from data that extend anisotropically to 5.2 Å, 5.2 Å and 3.7 Å resolution, in conjunction with previously determined structures of individual domains. The highly intertwined structure illustrates why chemical activity depends on formation of the synaptic complex, and reveals that individual domains have different roles when bound to different sites. The structure also provides explanations for the increased stability of the final product complex and for its preferential recognition by the ATP-dependent unfoldase ClpX. Although MuA and many other recombinases share a structurally conserved ‘DDE’ catalytic domain, comparisons among the limited set of available complex structures indicate that some conserved features, such as catalysis in trans and target DNA bending, arose through convergent evolution because they are important for function.

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Figure 1: Transposition pathway and structure determination.
Figure 2: Transpososome structure.
Figure 3: Stereo close-up view of interactions near the Mu DNA–target junction.
Figure 4: Model for a transpososome assembled on full left (reddish) and right (blue) bacteriophage ends.
Figure 5: Comparison of DDE recombinase–DNA complexes.

Accession codes

Primary accessions

Protein Data Bank

Data deposits

Coordinates and structure factors were deposited at the Protein Data Bank under accession 4fcy.

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Acknowledgements

We thank K. Mizuuchi for initiating this project, K. K. Swinger and B. Vertessy for early crystallization efforts, and X. Yang and the staff of APS beamlines 14, 19 and 21 for assistance with data collection. This work was funded in part by NIH grant GM086826 (to P.A.R.).

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S.P.M. carried out most of the crystallographic work, Y.Z.P. grew the first diffracting transpososome crystals and assisted with all other aspects of the project, and P.A.R. designed the project and assisted in computational work and interpretation of the results.

Corresponding author

Correspondence to Phoebe A. Rice.

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The authors declare no competing financial interests.

Supplementary information

Supplementary Information

This file contains Supplementary Figures 1-4, Supplementary References and Supplementary Table 1. (PDF 1162 kb)

Ribbon drawing of the transpososome structure rotating 360°

The complex is rotating about the crystallographic twofold axis that relates the red and blue halves. Colours are as in the main text: bacteriophage Mu end DNAs are red and blue, target DNA black, and the scissile phosphate and active site residues are yellow. The darker-colored subunits catalyze DNA cleavage and strand transfer and the lighter-colored subunits aid in complex assembly and stability. (MPG 4785 kb)

Closeup view of the experimental electron density, after improvement with Parrot, and contoured at 1.3 and 2.3 Sigma, rotating 360°

The rotation axis and colors are as in the main text and Supplementary Video 1. (MPG 9535 kb)

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Montaño, S., Pigli, Y. & Rice, P. The Mu transpososome structure sheds light on DDE recombinase evolution. Nature 491, 413–417 (2012). https://doi.org/10.1038/nature11602

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