Mechanism for loading the E. coli beta clamp on DNA

State University Of New York At Buffalo, Buffalo NY

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Abstract

The long term goal of this research is to understand how organisms coordinate the replication and repair of their genomes. The use of processivity clamps that must be loaded onto DNA during replication of chromosomes by an ATP dependent clamp loader complex is highly conserved by all forms of life. Understanding how organisms coordinate DNA replication and repair is the fundamental basis for understanding human diseases such as cancer. The goal of the work proposed here is to better define the mechanism of clamp loading in the E. coli model system. The E. coli clamp loader (y complex) is a multisubunit complex that undergoes a conformational change upon binding ATP which facilitates opening of the p clamp and its subsequent loading onto DNA. The contacts of the individual clamp loader subunits with P differ between the ATP bound and free states of y complex. The interactions of the y complex subunits with p in both the presence and absence of ATP will be determined. Biochemical assays using various clamp and clamp loader mutants will be uilized to accomplish this. The insight gained from E. coli can provide a framework for elucidating the mechanisms of eukaryotes, including humans.