Measurement of protein-protein interactions applied to protein crystallization in salt and polyethylene glycol solutions
Date
2005
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Publisher
University of Delaware
Abstract
Crystallization is currently the limiting step in determining the structure of proteins using x-ray diffraction techniques. The second osmotic virial coefficient is related to the crystallization conditions, and thus it is the parameter of choice for studying protein-protein interactions, but experimental difficulties limit the efficiency with which measurements can be performed. A recently developed method, self-interaction chromatography (SIC), allows rapid and efficient measurement of the second osmotic virial coefficient and consequently collection of data to develop further an understanding of protein-protein interactions. The main goal of this study is therefore to investigate protein-protein interactions by measuring the second osmotic virial coefficient in salt and polyethylene glycol solutions, so as to develop insights about the solution conditions that promote protein crystallization. ☐ The second osmotic virial coefficient was measured for ovalbumin at different pH values and in different salt solutions. Changes observed in the interaction trends around pH 4-5 show the importance of glutamic and aspartic acid in protein-protein interactions. Differences among salts, ranging in properties from chaotropic, which have very little effect on protein-protein interactions, to kosmotropic, which have a strong effect, are measured and explained in terms of their effects on protein hydration. Comparisons with other proteins, namely ribonuclease A, myoglobin, α- lactalbumin and BSA, show that these observations can be generalized. The Hofmeister series is discussed from the perspective of these results and Raman scattering measurements of salt solutions are used to show that the effects observed do not correspond to dramatic changes in the water structure. ☐ Protein-protein interactions were also investigated for ovalbumin, ribonuclease A, myoglobin and α-lactalbumin in solutions of polyethylene glycol of different molecular weights. The results are in agreement with a depletion mechanism, and the trends can be modeled using the classic Asakura-Oosawa potential. However, under certain conditions other mechanisms can be suggested. Protein crystallization in polyethylene glycol solutions is explained using these results, and the validity of the Asakura-Oosawa potential for proteins is discussed.