The overall goal of our work is to establish structure-function relationships in mutagenesis and DNA repair and to develop computational methods that can be used to predict patterns by which repair enzymes recognize damaged DNA. We propose to determine solution structures of several DNA molecules containing mutagenic adducts, the zinc finger motif of Fpg protein, and its 1:1 complex with DNA. Adducts to be studied include acetylaminofluorene-C(8)- and N2-guanine, aminofluorene-G(8)- guanine, phenilimidazopyridine-C(8)-guanine, 8-oxoguanine, 8-oxoadenine, 8-aminoguanine, as well as abasic sites. Based on mutagenesis studies conducted in (Project 2), these adducts will be incorporated into DNA duplexes as models for damaged DNA; misaligned bulged duplexes, as models for frameshift mutagenesis; and primer-templates as models for replication fork intermediates. These adducts will be synthesized in the laboratories of Dr. Johnson (Project i); several bases will be isotopically labeled to enhance NMR resolution. One- and two-dimensional high resolution NMR experiments will be performed on the adducted DNA molecules, using the 600 MHz spectrophotometer at Stony Brook. NMR data will be used to derive dihedral-, distance- and volume-restraints which, when incorporated into molecular mechanics and dynamics calculations, will establish the solution structure of the above-mentioned adducts. Calculations will run on our Silicon Graphics computers, as well as on NSF supercomputers. We propose to expand our computational methods for macromolecular docking based on hydrogen bond pattern recognition by including dynamic flexibility and water bridging to our current, rigid- body molecular model. Enhancements of our computer programs will be coded and run on the hypercube parallel computer facility at the Applied Mathematics Department.
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