The DNA architectural protein Xis regulates the construction of higher-order nucleoprotein intasomes that integrate and excise the genome of phage lambda from the Escherichia coli chromosome. Xis modulates the directionality of site-specific recombination by stimulating phage excision 10(6)-fold, while simultaneously inhibiting phage reintegration. Control is exerted by cooperatively assembling onto a approximately 35-bp DNA regulatory element, which it distorts to preferentially stabilize an excisive intasome. Here, we report the 2.6-A crystal structure of the complex between three cooperatively bound Xis proteins and a 33-bp DNA containing the regulatory element. Xis binds DNA in a head-to-tail orientation to generate a micronucleoprotein filament. Although each protomer is anchored to the duplex by a similar set of nonbase specific contacts, malleable protein-DNA interactions enable binding to sites that differ in nucleotide sequence. Proteins at the ends of the duplex sequence specifically recognize similar binding sites and participate in cooperative binding via protein-protein interactions with a bridging Xis protomer that is bound in a less specific manner. Formation of this polymer introduces approximately 72 degrees of curvature into the DNA with slight positive writhe, which functions to connect disparate segments of DNA bridged by integrase within the excisive intasome.