The Molecular Mechanisms of the Mre11/Rad50 DNA Repair Complex

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Intellectual Merit: The goal of this project is to achieve a mechanistic understanding of the Mre11/Rad50 (MR) DNA repair complex, which is involved in the processing of DNA double-strand breaks (DSBs) within eukaryotes, archaea, and some bacteriophage. DSBs are among the most deleterious forms of DNA damage and their improper repair may cause gross chromosomal rearrangements, leading to cellular dysfunction or death. The dual activities of the MR complex (ATP hydrolysis and nuclease) are required for the repair of DSBs that have been chemically modified by toxins or proteins. This research project will focus on defining, in quantitative terms, the kinetics, thermodynamics, and conformational dynamics of the MR complex. The objectives of this research project are to: 1) Define the functional role and mechanism of ATP binding and hydrolysis by Rad50; 2) Determine the mechanism behind the metal-dependent change in MR complex nuclease activity and to discover the possible roles of additional proteins in DSB resection; 3) Characterize the conformational changes in Mre11 and determine the role of Mre11 dimerization. The completion of this project is expected to increase understanding of DSB repair and, more generally, how organisms maintain their genomic integrity through multiple generations. Broader Impacts: This project involves collaborations with both genetic and structural biology laboratories and will therefore expose and educate undergraduate and graduate students to the synergy that results from the combination of biochemical, biophysical, and genetic research. The project will provide high quality training opportunities for high school, undergraduate, and graduate students. In addition to the multi-year training of students in the PI's home department, the PI participates in the Women in Science and Engineering internship program and will become a mentor for the George Washington Carver internship program. The goal of these programs are to increase the diversity of talented individuals choosing science as a career. The PI is also developing a course that places science journalism students in a research laboratory where they will attend group meetings, interview members of the laboratory, and observe daily activities. The objective is for the journalism students to appreciate the manner and pace at which experimental findings are obtained. The laboratory members who are being interviewed and observed will also benefit by learning to explain their research to non-experts. In addition, the PI is developing a course that combines enzyme theory, practical enzyme characterization, and modern data fitting/model discrimination analysis. Throughout the course the PI will use examples from his own research, and when possible, the students that generated the data under discussion will present the experiments to their peers. This type of peer-to-peer teaching has been found to be a very effective learning tool for both the peer-teachers and peer-students.This project is co-funded by the Genetic Mechanisms Cluster in the Division of Molecular and Cellular Biosciences and by the Experimental Program to Stimulate Competitive Research.