Intellectual Merit: Understanding the biochemical pathways associated with how genes and genomes are inherited, and changed, remains an urgent priority. The goal of this work is to gain structure-based understanding of one such pathway in bacteria, e.g., nucleotide excision repair (NER) of DNA damage induced by exposure to ultraviolet light. The first steps of bacterial NER are performed by three proteins, UvrA, UvrB, and UvrC, which associate into several dynamic multi-protein complexes as repair proceeds. The UvrA-UvrB ensemble scans the genome, distinguishing lesion-containing damaged DNA from native, and, then, cooperates with the UvrC nuclease and other factors to repair the damage. This work will test novel hypotheses for three incompletely understood mechanisms of NER: a) discrimination of native from damaged, b) ATP/ADP dynamics during NER, and c) localization of UvrB to the lesion. This study takes a multi-disciplinary approach, integrating structural, biochemical, single-molecule, and biophysical strategies to understand relationships between the structure, function, and dynamics of relevant macromolecules as the genome is scanned for lesions. Structure-based analyses of genetic mechanisms during NER will provide deeper insights into the connections between DNA repair, DNA replication, inheritance, and transcription. More broadly, analysis of the relationship between shape-based recognition of DNA, proposed by the PI as a mechanism for discriminating damaged and native DNA, and protein conformational changes will contribute to a comprehensive understanding of fundamental principles that underlie the way that DNA is recognized by proteins in biological systems.
Broader Impacts: One of the central challenges for science education is how to impart quantitative skills to younger colleagues. The PI approaches this problem by using research on DNA repair, a fundamental biological process, as a platform to teach such skills. This project builds on the PI's past efforts at Harvard University, which now continue at City College of New York. CCNY is a public Ph.D.-granting institution that attracts undergraduate/doctoral students from diverse socioeconomic strata, and ethnic and racial backgrounds. Towards advancing education in science, technology, engineering and mathematics (STEM), the PI will teach two courses at CCNY, into which the described research will be integrated, and which will impact ~100 undergraduate/doctoral students. In addition, in partnership with The Urban Assembly, the PI will perform educational outreach in the New York Public School system, which will include meetings with students (grades 6-12), and collaborations with teachers on lesson plans. The science performed in the PI's group is multi-disciplinary and involves close collaborations between several research groups. The PI actively mentors post-docs, and, they, in turn, teach and mentor junior colleagues. The results of the research will be published in peer-reviewed journals. Research materials will be made available to the scientific community in several ways, including depositions to the Protein Data Bank.
