Abstract
Recombination directionality factors (RDFs), or excisionases, are essential players of prophage excisive recombination. Despite the essentially catalytic role of the integrase in both integrative and excisive recombination, RDFs are required to direct the reaction towards excision and to prevent re-integration of the prophage genome when entering a lytic cycle. KplE1, HK620 and numerous (pro)phages that integrate at the same site in enterobacteria genomes (such as the argW tRNA gene) all share a highly conserved recombination module. This module comprises the attL and attR recombination sites and the RDF and integrase genes. The KplE1 RDF was named TorI after its initial identification as a negative regulator of the tor operon. However, it was characterized as an essential factor of excisive recombination. In this study, we designed an extensive random mutagenesis protocol of the torI gene and identified key residues involved in both functions of the TorI protein. We show that, in addition to TorI-TorR protein-protein interaction, TorI interacts in solution with the IntS integrase. Moreover, in vitro, TorR and IntS appear to compete for TorI binding. Finally, our mutagenesis results suggest that the C-terminal part of the TorI protein is dedicated to protein-protein interactions with both proteins TorR and IntS.
Highlights
The name bacteriophage encompasses all bacterial viruses, including temperate phages which have the particularity to integrate their genomes into their hosts, becoming prophages
Whereas host factors can modulate the efficiency of the integrase mediated reactions, most of the time directionality is driven by recombination directionality factors (RDF) or excisionases [9,10,11,12]
In order to identify critical residues involved in the anti-response regulator (anti-RR) and/or the excisive recombination activity of the TorI RDF, we designed a tester strain that can report both activities
Summary
The name bacteriophage encompasses all bacterial viruses, including temperate phages which have the particularity to integrate their genomes into their hosts, becoming prophages. Constitutes a key step in lysogenic development since it is required for integration as well as for excision of the prophage genome [6] This reaction is mediated in both directions by a specific recombinase, called integrase, that belongs either to the tyrosine or the serine recombinase families [7,8]. We studied one of them in particular, the KplE1 (or CPS53) prophage This latter is inserted into the argW tRNA gene at 2,474 kb on the E. coli chromosome and contains 16 ORFs. Most have unknown functions, whereas we previously characterized the role of the first gene intS and the last one torI in site-specific recombination [20,21]. We designed an extensive random mutagenesis protocol of the torI gene to identify critical residues involved in the anti-response regulator (anti-RR) and/or the excisive recombination activity functions of the TorI protein
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
