This research is designed to elucidate the molecular mechanisms of mutagenesis using the well-studied bacterium, Escherichia coli. It is assumed that mechanisms characterized in this and other studies will be applicable to all organisms including humans where mutational events are known to give rise to genetic diseases and various forms of cancer. Spontaneous mutations result from several cellular processes including DNA replication which is one area of emphasis in this study. Mutant E. coli strains, called mutators, are available which are known to be defective in some aspect in the fidelity of DNA replication. By constructing double mutator strains it is possible to study the interactions between two different systems, both involved in providing fidelity. Spontaneous mutagenesis will also be studied by analyzing putative antimutator alleles that have recently been isolated. The existence of an antimutator allele must define a locus that makes a significant contribution toward the production of spontaneous mutations. The first approaches to be addressed involve the mapping of the putative antimutators and the characterization of their normal gene function(s). Oxidative DNA damage from active oxygen species has been hypothesized to play a crucial role in mutagenesis as well as other biological processes such as aging and carcinogenesis. Until recently studies on the role of oxygen in spontaneous mutagenesis have been hampered by the inability to conduct experiments in an environment totally free of oxygen. Anaerobic chambers are now available that allow mutagenesis experiments to be carried out in an oxygen-free environment. Mutation frequencies and mutational specificity will be determined under anaerobic conditions and then compared with aerobic mutagenesis, thus demonstrating the possible role of oxygen in mutagenesis.
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