The stereochemistry and thermodynamics of DNA-target recognition by specific DNA-binding proteins will be studied. The model system used is the E2 transcriptional regulatory protein and replication factor of the cancer- associated papillomaviruses. The techniques used will include X-ray crystallography, titration calorimetry, quantitative gel-retardation assays and mutational analyses of the proteins. The following specific goals will be pursued: To elucidate the structural basis for DNA target discrimination by the E2 protein of the human papillomaviruses. To establish the energetics of specific DNA recognition by the E2 proteins. To determine the role of the 'hinge' region of the E2 repressor proteins in DNA-binding. To determine the three-dimensional structure of the entire B2 protein and its complex with DNA. This study will go beyond the cataloguing of specifying intermolecular contacts. It will explicitly define and quantitate the contributions of indirect factors such as intrinsic deformability and changes in solvent accessible surface areas in mediating the DNA target selectivity exhibited by the E2 proteins. Insights regarding the contributions to specificity and affinity of these features will then be extrapolated to other protein-DNA systems. The detailed structural information will provide a basis for the rational design of molecules that could modulate E2-DNA complex formation. Such agents would be of therapeutic value in the treatment of papillomavirus-caused pre-cancerous lesions.
