Our long-range goal is to achieve a detailed understanding of the molecular mechanisms involved in site-specific recombination. As a model we are studying the Int-dependent recombination pathway of the lambdoid phages, lambda, 080 and P22. The major portion of our effort will continue to focus on the lambda system; a great deal is known about this system and the rate of progress in its analysis has recently been taking some dramatic steps forward. The experiments in this application are intended to complement studies being carried out in other laboratories. For the purpose of presenting our research plan, the aims of this project can be viewed as addressing seven specific questions or problems. The first five are concerned primarily with the general mechanisms of site-specific recombination and the last two concentrate specifically on the remarkable control over directionality in this genetic transaction. 1. What are the limitations on the size of the overlap region? 2. Are the differences (in DNA sequence and affinity for Int) among the four core-type sites an intrinsic feature of the recombination reaction? 3. How do the arm-type Int binding sites function during recombination? 4. What additional insights into the basic mechanisms of site-specific recombination are provided by the features of the 080 and P22 recombination systems? 5. To use gel electrophoresis mobility shifts and electron micro-scopy to study the nature of att site complexes and the kinds of interactions that determine integrative and excisive recombination. 6. What is the minimal set of att site elements necessary for excisive recombination in comparison to the set defined for integrative recombination? 7. What are the roles of the individual arm-type binding sites as (potential) determinants in the control of directionality?
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