STRUCTURE OF THE HOLLIDAY JUNCTION INTERMEDIATE IN CRE-LOXP SITE-SPECIFIC RECOMBINATION

We have determined the X-ray crystal structures of two DNA Holliday ju nctions (HJs) bound by Cre recombinase, The HJ is a four-way branched structure that occurs as an intermediate in genetic recombination path ways, including site-specific recombination by the lambda-integrase fa mily. Cre recombinase is an integrase family member that recombines 34 bp loxP sites in the absence of accessory proteins or auxiliary DNA s equences. The 2.7 Angstrom structure of Cre recombinase bound to an im mobile HJ and the 2.5 Angstrom structure of Cre recombinase bound to a symmetric, nicked HJ reveal a nearly planar, twofold-symmetric DNA in termediate that shares features with both the stacked-X and the square conformations of the HJ that exist in the unbound state. The structur es support a protein-mediated crossover isomerization of the junction that acts as the switch responsible for activation and deactivation of recombinase active sites. In this model, a subtle isomerization of th e Cre recombinase-HJ quaternary structure dictates which strands are c leaved during resolution of the junction via a mechanism that involves neither branch migration nor helical restacking.