Identification of the lambda integrase surface that interacts with Xis reveals a residue that is also critical for Int dimer formation.

Abstract

Lambda integrase (Int) is a heterobivalent DNA-binding protein that together with the accessory DNA-bending proteins IHF, Fis, and Xis, forms the higher-order protein-DNA complexes that execute integrative and excisive recombination at specific loci on the chromosomes of phage lambda and its Escherichia coli host. The large carboxyl-terminal domain of Int is responsible for binding to core-type DNA sites and catalysis of DNA cleavage and ligation reactions. The small amino-terminal domain (residues 1-70), which specifies binding to arm-type DNA sites distant from the regions of strand exchange, consists of a three-stranded beta-sheet, proposed to recognize the cognate DNA site, and an alpha-helix. We report here that a site on this alpha-helix is critical for both homomeric interactions between Int protomers and heteromeric interactions with Xis. The mutant E47A, which was identified by alanine-scanning mutagenesis, abolishes interactions between Int and Xis bound at adjacent binding sites and reduces interactions between Int protomers bound at adjacent arm-type sites. Concomitantly, this residue is essential for excisive recombination and contributes to the efficiency of the integrative reaction. NMR titration data with a peptide corresponding to Xis residues 57-69 strongly suggest that the carboxyl-terminal tail of Xis and the alpha-helix of the aminoterminal domain of Int comprise the primary interaction surface for these two proteins. The use of a common site on lambda Int for both homotypic and heterotypic interactions fits well with the complex regulatory patterns associated with this site-specific recombination reaction.