The site-specific recombinase (Int) of bacteriophage is a heterobivalent DNA binding protein and is comprised of three domains: an N-terminal domain that binds with high affinity to “arm-type” sequences within the recombination target DNA (att sites), a C-terminal domain that contains all of the catalytic functions, and a central domain that contributes significantly to DNA binding at the “core-type” sequences where DNA cleavage and ligation are executed. To further analyze the interaction between Int and core-type DNA sequences, we constructed a family of core-type DNA oligonucleotides, each of which contained the photo-reactive analog 4-thio-deoxythymidine (4-thioT) at a different position. When tested for their respective abilities to promote covalent crosslinks with Int after irradiation with UV light at 366nm, one oligonucleotide stood out dramatically. The 4-thioT substitution on the DNA strand opposite the site of Int cleavage led to photo-induced crosslinking efficiencies of approximately 20%. The efficiency and specificity of Int binding and cleavage at this 4-thioT substituted core site was shown to be largely uncompromised and its ability to participate in a full site-specific recombination reaction was reduced only slightly. Identification of the photo-crosslinked residue as Lys141 in the central domain provides, along with other results, several insights about the nature of core-type DNA recognition by the bivalent recombinases of the -Int family. The high efficiency of photo-crosslinking at this position also affords a useful handle for dissecting the higher-order organization of the structures associated with this recombination pathway.