Quantifying Single Molecule DNA-ligand Interactions

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DNA interactions are critical aspects of the life cycle of the cell, but the mechanisms that govern many of these interactions are not well understood at the molecular level. There are many examples of important DNA interactions that are difficult to study using traditional biophysical methods. For these cases the investigators develop new methods in which small forces are applied to stretch single DNA molecules in the presence of proteins and small molecules. By quantifying force-dependent DNA interactions, the investigators can determine the molecular properties that regulate these interactions. The results from these measurements provide unique insights into important biological processes. The project also involves training and professional development of undergraduate and graduate students from a variety of backgrounds. This includes full-time undergraduates working in the lab as part of Northeastern's co-op program, scientific training at a women's college, and providing opportunities for students from a regional university with a majority of first-generation college students and a high percentage of low-income students. This project uses DNA stretching with optical tweezers to probe biologically important molecular-level nucleic acid interactions that are particularly well-suited to probe with single molecule methods. Small molecule-DNA interactions will be studied, particularly for cases in which significant DNA structural rearrangements are required for DNA binding and dissociation. The properties of these small molecules that determine the DNA unfolding landscape will be measured, revealing critical information needed for the design of useful new ligands. LINE1 is a retrotransposon that is active in human cells. Recent studies probed the interaction between an essential LINE1-encoded protein, ORF1p, with nucleic acid, showing that its universally conserved coiled coil motif facilitates ORF1p oligomerization on nucleic acid. This project will probe the biophysical mechanism responsible for oligomerization, a process that is essential for retrotransposition. Finally, the activity of several E. coli DNA polymerases and polymerase manager or accessory proteins will be studied. These studies will reveal detailed interactions that are part of the bacterial response to DNA damage, a response essential for all life. This project is jointly funded by the Molecular Biophysics Cluster in the Division of Molecular and Cellular Biosciences and the Physics of Living Systems Program in the Division of Physics.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.