The protein-protein interactions required for assembly of the Tn3 resolution synapse

Rowland, S.-J., Boocock, M. R., Burke, M. E., Rice, P. A. and Stark, W. M. (2020) The protein-protein interactions required for assembly of the Tn3 resolution synapse. Molecular Microbiology, 114(6), pp. 952-965. (doi: 10.1111/mmi.14579) (PMID:33405333)

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Abstract

The site‐specific recombinase Tn3 resolvase initiates DNA strand exchange when two res recombination sites and six resolvase dimers interact to form a synapse. The detailed architecture of this intricate recombination machine remains unclear. We have clarified which of the potential dimer‐dimer interactions are required for synapsis and recombination, using a novel complementation strategy that exploits a previously uncharacterized resolvase from Bartonella bacilliformis (“Bart”). Tn3 and Bart resolvases recognize different DNA motifs, via diverged C‐terminal domains (CTDs). They also differ substantially at N‐terminal domain (NTD) surfaces involved in dimerization and synapse assembly. We designed NTD‐CTD hybrid proteins, and hybrid res sites containing both Tn3 and Bart dimer binding sites. Using these components in in vivo assays, we demonstrate that productive synapsis requires a specific “R ” interface involving resolvase NTDs at all three dimer‐binding sites in res . Synapses containing mixtures of wild‐type Tn3 and Bart resolvase NTD dimers are recombination‐defective, but activity can be restored by replacing patches of Tn3 resolvase R interface residues with Bart residues, or vice versa . We conclude that the Tn3 /Bart family synapse is assembled exclusively by R interactions between resolvase dimers, except for the one special dimer‐dimer interaction required for catalysis.

Item Type:Articles
Additional Information:This work was funded by Wellcome Trust grant 072552.
Status:Published
Refereed:Yes
Glasgow Author(s) Enlighten ID:Stark, Professor Marshall and Boocock, Dr Martin and Rowland, Dr Sally-Jane
Authors: Rowland, S.-J., Boocock, M. R., Burke, M. E., Rice, P. A., and Stark, W. M.
College/School:College of Medical Veterinary and Life Sciences > Institute of Molecular Cell and Systems Biology
Journal Name:Molecular Microbiology
Publisher:Wiley
ISSN:0950-382X
ISSN (Online):1365-2958
Published Online:29 July 2020
Copyright Holders:Copyright © 2020 The Authors
First Published:First published in Molecular Microbiology 114(6): 952-965
Publisher Policy:Reproduced under a Creative Commons License

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