1. The protein–protein interactions required for assembly of the Tn3 resolution synapse.
- Author
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Rowland, Sally‐J., Boocock, Martin R., Burke, Mary E., Rice, Phoebe A., and Stark, W. Marshall
- Subjects
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PROTEIN-protein interactions , *SYNAPSES , *BINDING sites , *GENETIC recombination , *RECOMBINASES - 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. [ABSTRACT FROM AUTHOR]
- Published
- 2020
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