ENZYMOLOGICAL AND GENETIC STUDIES OF DNA POLYMERASES

University Of California Berkeley, Berkeley CA

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Abstract

The major focus of the work will be on the application of topological and genetic approaches to the study of several key enzymes in DNA metabolism. This will involve developing new techniques for determining DNA structure and expanding the theory of DNA folding. In addition, we hope to isolate mutants of E. coli topoisomerase III to determine its function in vivo. We will also investigate the enzymes in E. coli that metabolize DNA knots and catenases and measure the functional level of DNA supercoiling in this organism and perhaps in yeast. Using a rigorous topological method, the mechanism of chromosome segregation in several organisms will be tested. Analogous methods will be brought to bear on the mechanism of topoisomerases. We will continue our studies of transcription by RNA polymerase III and its accessory factors. This will involve purification of the factors and determination of their role, analysis of the formation of transcription complexes, measurement of the DNA binding sites, and exploration of the striking increase in a polymerase III transcript after neoplastic transformation.