Macromolecular crowding in vitro and in cells

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Protein-protein interactions regulate all cellular functions, yet they are almost always studied in water rather than in the complex and crowded environment that exists inside cells. The results of this project will fill a key gap that prevents a complete description of metabolism. The goal of moving quantitative biophysics from simple solutions to crowded environments, including inside living cells, is a major challenge with important outcomes. The knowledge gained will add to both the fundamental understanding of biology and inform efforts to produce and stabilize protein-based reagents. The work will also facilitate the training of undergraduate and graduate students in the practice of cutting-edge research. All these efforts are key to building the US bioeconomy.The overall goal of the project is to gain broadly-applicable knowledge about protein-protein interactions under physiologically-relevant conditions. The focus is on homodimeric interactions in vitro, in Escherichia coli cells and in zebrafish (Danio rerio) oocytes. The overarching hypothesis is that electrostatic interactions between cosolutes and protein complexes dominate the effect of hard-core excluded volume under physiologically-relevant conditions. More specifically, electrostatic attractions between cosolutes and protein complexes are expected to decrease stability of the complexes, whereas electrostatic repulsions are expected to increase stability. The supported research will also deliver to modelers and theoreticians the information required to produce accurate simulations of cellular processes. This project is supported by the Molecular Biophysics Cluster of the Molecular and Cellular Biosciences Division in the Biological Sciences Directorate.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.