MECHANISMS OF FIDELITY AND MUTAGENESIS

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The broad long-term goal of this project is to understand how mutations occur from the replication of carcinogen-modified DNA. The objective of this grant proposal is to elucidate the interactions between DNA polymerase and the minor groove of DNA. The specific aims are to determine which sites in the minor groove of the DNA interact with the DNA polymerase during replication. These interactions will be examined during the replication of the normal DNA bases and theta6-methylguanine, a carcinogen- modified base. Differences observed between correct and incorrect replication will give clues as to the mechanisms that polymerases use to replicate DNA with high fidelity. The health-relatedness of the project is in carcinogenesis. Mutations that occur during replication of normal DNA and carcinogen-modified DNA can lead to cancer. Understanding the mechanism of the formation of mutations might alloW us to predict which compounds are mutagens. The research design is to synthesize DNA analogs in which sites in the minor groove are altered so that they can not participate in hydrogen bonds. The kinetics of incorporation of modified nucleotide triphosphates opposite modified DNA will be measured. Large reduction in rates of replication will indicate that the altered site was involved in a hydrogen bond with the polymerase.