Abstract

   We use homology modeling, molecular dynamics simulations,  and comprehensive sequence comparison methods to determine how DNA repair enzymes  recognize subtle differences in the structure and dynamics of DNA sites  damaged by low dose ionizing radiation (IR). Our combined computational  and experimental approach focuses on the apurinic/apyrimidinic endonuclease  1 (APE1) to elucidate the roles of residues close to the DNA sites or that  mediate interactions with other enzymes in the base excision repair (BER) pathway. Our studies are based on our expert system MASIA for analysis of multiple aligned sequences and 3D structure prediction, recent high-resolution X-ray crystal structures of APE1-DNA complexes, and functional studies of human wild-type and mutant APE1. We use simulations to analyze global conformational changes of APE1-DNA complexes and the flexibility of functionally important residues of human APE-1. Structural and functional hypotheses based on these calculations are tested using functional assays of mutant APE-1. The capabilities of MASIA are expanded to include a new method for numerical description of the chemical state of each column of a protein sequence matrix that are used to further analyze motifs in the bacterial and mammalian members of the family. We derive new sequence motifs from the gene and protein sequences of the currently known APE1 family for specific aspects, such as primary DNA recognition, catalysis, and protein-protein interaction. These motifs are tested by determining the activities of mutated APE1 proteins in in vivo and in vitro assays. Screening gene databases with these novel motifs revealed new candidates for APE related proteins.