Probe the conformational dynamics of a protein targeting machine at single molecule resolution

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Observing the dynamic movements of molecular machines in action and deciphering their roles in biological function are the frontiers of life science research. This project aims to elucidate the molecular movements in a protein targeting machine, the signal recognition particle (SRP), as it delivers ~30% of newly synthesized proteins to the correct biological membrane in the cell. The project will generate valuable reagents, tools, and assays that are useful to many other researchers. The results of the research will be disseminated through publications in academic journals as well as animated movie illustrations that will be available to the general public. The education component of the proposal will emphasize the training of graduate students and postdoctoral scholars in multidisciplinary biochemical and biophysical research. It will also expose high school and undergraduate students to state-of-the-art research tools and provide leadership experience for the graduate students and postdocs.More specifically, this project will combine the expertise of the Shan lab in mechanistic biochemistry and the Weiss lab in biophysics to decipher the dynamic motions that drive co-translational protein targeting by SRP at single molecule resolution. The recent works by the Shan lab indicated the presence of multiple largescale conformational rearrangements in SRP during the protein targeting cycle. This project will develop solution-based single molecule fluorescence assays to directly observe the conformational rearrangements in SRP during targeting, decipher the molecular forces that drive them, and understand their regulation by the GTPase cycle of SRP and by spatial and temporal signals in the pathway. The results will elucidate the molecular basis for the spatiotemporal control of this targeting machine, and reveal generalizable principles that underlie the action of nucleotide-driven macromolecular machines in biology.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.