Kinetically-stabilized mesoscopic protein aggregates

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It is generally accepted that a protein's amino acid sequence is optimized so that the molecule consistently folds into a unique, stable 3D shape while avoiding aggregation with other biomolecules. Yet this view is overly simplistic. On the one hand, many proteins are not monolithic structures; for instance, myoglobin must reliably deform to let in oxygen. On the other hand, under certain conditions, proteins may aggregate into amyloid fibers while in a misfolded conformation. Although very complex, these phenomena can still be understood thermodynamically. Proteins are deformable given a sufficiently large native state entropy, while amyloid fibers are stabilized by multiple amino acid contacts within the fiber. Much less understood are the mesoscopic liquid-like aggregates recently found in several protein solutions, including lysozyme, hemoglobin, and gamma-crystallin. These clusters are important as nucleation sites for protein crystals. The objective of this project is to test a microscopic hypothesis for the formation of the mesoscopic clusters by which the clusters result from the formation of long-lived protein complexes stabilized at high densities. The unusually large cluster size stems from the long lifetime of the complexes, which in turn owe their stability to the conformational freedom of constituent protein molecules. A key component of the proposed mechanism is that proteins partially unfold prior to binding and may even undergo domain swapping, whereby protein segments bind to complimentary segments on a different protein. The PIs will employ a combination of dynamic light scattering, Brownian microscopy, amide exchange NMR, and theoretical modeling to test this mechanism in several systems, including proteins barnase, RNase A, hemoglobin, insulin, and lysozyme.This research is a multidisciplinary effort by a theoretical physical chemist and experimental chemical engineer. A synthesis of a wide range of physicochemical and biochemical measurements and physical modeling will be undertaken to tackle this difficult problem; the outcome is expected to produce fundamental understanding of biological processes and provide new avenues for making new materials. Graduate, undergraduate, and high school students will be trained at the interface between physics, chemistry, and biology. The home institution of the PIs, the University of Houston, is one of the most ethnically diverse research universities in the nation, and is one of only three Hispanic-Serving Institutions in the US. Existing collaborations with local writers and radio personalities will be utilized to publicize the societal benefits of the research. This project is jointly supported by the Molecular Biophysics Cluster in the Division of Molecular and Cellular Biosciences and the Physics of Living Systems Program in the Physics Division.