Cleavage of the site-specific recombination protein gamma delta resolvase: the smaller of two fragments binds DNA specifically.

Abstract

The transposon Tn3-encoded 20,500-dalton .gamma..delta. resolvase monomer was cleaved by chymotrypsin into a 5000-dalton COOH-terminal fragment and a 15,500-dalton NH2-terminal fragment that were purified. Two crystal forms of the large fragment were obtained, 1 of which is isomorphous with crystals of the native protein, showing that the large fragment makes the protein-protein contacts in the crystal and that the small fragment is segmentally disordered relative to the large fragment. Nuclease protection demonstrated that the small fragment binds specifically to all 3 DNA binding sites protected by resolvase. Unlike native resolvase, which binds to all 3 complete sites with equal affinity, the small fragment binds to each of the 6 half sites with a different affinity. It was not possible to demonstrate specific DNA binding of the larger fragment. Thus, resolvase has a modular construction analogous to that found for some repressors and activators; its COOH-terminal domain recognizes specific sequences in the DNA and its NH2-terminal domain mediates protein-protein interactions and probably has the enzymatic activity.