To map the protein-protein and protein-DNA interactions involved in λ site-specific recombination, Int cleavage assays with suicide substrates, nuclease protection patterns, gel retardation experiments, and quantitative Western blotting were applied to wild-type attL and attL mutants. The results lead to a model in which one IHF molecule bends the attL DNA and forms a higher order complex with the three bivalent Int molecules required for excisive recombination. It is proposed that each of the Int molecules binds in a unique manner: one bridges two DNA binding sites in cis, one is held via its high affinity amino-terminal DNA binding domain, and the third depends upon protein-protein interactions in addition to its low affinity carboxy-terminal DNA binding domain. This protein-DNA complex contains two unsatisfied DNA binding domains, each with a different sequence specificity, and is well suited to specific interactions with an appropriate recombination partner.
Bibliographical noteFunding Information:
We are greatly indebted to Noaman Hasan for the antibody against Int; David Brautigan and Carol Shriner for advice on immunostaining; Bonnie Tracy, Lucy Rodrigues, and Tina Oliveira for technical assistance: and Joan Boyles for preparation of the manuscript. This work was supported by National Institutes of Health grants GM33928 and All3544 and March of Dimes Basic Research Grant l-543.
All Science Journal Classification (ASJC) codes
- Biochemistry, Genetics and Molecular Biology(all)