ACTION OF SITE-SPECIFIC RECOMBINASES XERC AND XERD ON TETHERED HOLLIDAY JUNCTIONS

In Xer site-specific recombination, two related recombinases, XerC and XerD, mediate the formation of recombinant products using Holliday ju nction-containing DNA molecules as reaction intermediates. Each recomb inase catalyses the exchange of one pair of specific strands. By using synthetic Holliday junction-containing recombination substrates in wh ich two of the four arms are tethered in an antiparallel configuration by a nine thymine oligonucleotide, we show that XerD catalyses effici ent strand exchange only when its substrate strands are 'crossed'. Xer C also catalyses very efficient strand exchange when its substrate str ands are 'crossed', though it also appears to be able to mediate stran d exchange when its substrate strands are 'continuous'. By using chemi cal probes of Holliday junction structure in the presence and absence of bound recombinases, we show that recombinase binding induces unstac king of the bases in the centre of the recombination site, indicating that the junction branch point is positioned there and is distorted as a consequence of recombinase binding.