Extreme C-terminal element of SprA serine integrase is a potential component of the molecular toggle switch which controls the recombination and its directionality

In Bacillus subtilis, a sporulation-related gene, spsM, is disrupted by SP beta prophage, but reconstituted during sporulation through SP beta excision. The spsM reconstitution is catalyzed by a site-specific DNA recombinase, SprA, and its cognate recombination directionality factor, SprB. SprB interacts with SprA, directing the SprA-mediated recombination reaction from integration to excision; however, the details of the directionality control remains unclear. Here, we demonstrate the importance of the extreme C-terminal region (ECT) of SprA in the DNA recombination and directionality control. We created a series of SprA C-terminal deletants and examined their DNA-binding and recombination activities. Deletions in the ECT caused a loss of integration and excision activity, the magnitudes of which positively correlated with the deletion size. Gel shift study revealed that the loss of the integration activity was attributable to the failure of synaptic complex formation. The excision deficiency was caused by defective interaction with SprB. Moreover, alanine scanning analysis revealed that Phe532 is essential to interact with SprB. SprA(F532A), therefore, showed almost no excision activity, while retaining the integration activity. Collectively, these results suggest that the ECT plays the crucial roles in the interaction of SprA with SprB and possibly in the directional control of the recombination.

Automated summary

This study demonstrates the importance of the extreme C-terminal region (ECT) of SprA in DNA recombination and directionality control in Bacillus subtilis. The interaction between SprA and SprB is influenced by the alanine residue Phe532 in the extreme C-terminal region, affecting the directional control of the recombination reaction.