Site-specific recombination at dif by Haemophilus influenzae XerC

Abstract

Xer site-specific recombination at the Escherichia coli chromosomal site dif converts chromosomal dimers to monomers, thereby allowing chromosome segregation during cell division. dif is located in the replication terminus region and binds the E. coli site-specific recombinases EcoXerC and EcoXerD. The Haemophilus influenzae Xer homologues, HinXerC and HinXerD, bind E. coli dif and exchange strands of dif Holliday junctions in vitro. Supercoiled dif sites are not recombined by EcoXerC and EcoXerD in vitro, possibly as a consequence of a regulatory process, which ensures that in vivo recombination at dif is confined to cells that can initiate cell division and contain dimeric chromosomes. In contrast, the combined action of HinXerC and EcoXerD supports in vitro recombination between supercoiled dif sites, thereby overcoming the barrier to dif recombination exhibited by EcoXerC and EcoXerD. The recombination products are catenated and knotted molecules, consistent with recombination occurring within synaptic complexes that have entrapped variable numbers of negative supercoils. Use of catalytically inactive recombinases provides support for a recombination pathway in which HinXerC-mediated strand exchange between directly repeated duplex dif sites generates a Holliday junction intermediate that is resolved by EcoXerD to catenated products. These can undergo a second recombination reaction to generate odd-noded knots.