Separating proteins in aqueous solutions through fractional precipitation by addition of salts is one of the oldest protein separation methods, and it remains the only purification step for some industrial enzymes and an initial purification step for higher-value therapeutic proteins. However, despite its widespread application, the fundamental characteristics of protein precipitation by salt are still incompletely understood. The primary focus of this thesis is the experimental exploration of the phase behavior of single and binary protein systems during protein precipitation in concentrated ammonium sulfate and sodium chloride solutions, and the effect of solution conditions on protein interactions. Protein precipitation phase behavior of single protein systems was measured as a function of solution conditions for lysozyme and ovalbumin. X-ray powder diffraction (XRD) and optical microscopy were used to characterize the structure of the protein precipitates. These studies showed that crystallization after initial precipitation is feasible even at high ionic strengths if sufficient mixing time is allowed, and such behavior may be a more general phenomenon although different crystallization pathways may be found in some systems and under different conditions; for instance, different pathways are seen in ovalbumin-ammonium sulfate systems as a function of pH. A more important development is the interpretation that emerges of precipitation as a manifestation of a metastable liquid-liquid separation, with the precipitate being a kinetically trapped dense liquid phase. Furthermore, the diversity of possible dense phases (amorphous precipitate, crystal phases or gel) associated with liquid-liquid phase separation shows that understanding the effects of protein interactions of different kinds and at different ranges could have important consequences for the systematic control of protein phase behavior. The phase behavior in selective precipitation of lysozyme-ovalbumin binary mixtures was investigated at various conditions. The best combination of selectivity and recovery of lysozyme was achieved in pH 7 NaCl solutions. At an initial concentration of 30 mg/g H 2 O of lysozyme and 30 mg/g H 2 O of ovalbumin, more than 80% of the lysozyme was collected in precipitate form with 93% purity at ionic strengths above 3 m, while ovalbumin was collected in the supernatant at 80% purity. The purities of both proteins were therefore improved by at least 30% in one step. Moreover, crystallization following initial precipitation was observed in some binary precipitation experiments and indicates that binary precipitation, like single protein precipitation, is a metastable equilibrium state. The observations of selective precipitation behavior were correlated with values of the osmotic second virial coefficient, measured by cross-interaction chromatography (CIC). The results show that interaction measures such as protein self- and cross-interactions can be used as a predictor for general trends and/or inconsistencies in the separation behavior and for optimizing working conditions. Such a mechanistic relationship can obviate the need for extensive trial-and-error methods in order to optimize conditions for protein separations.

Because of the ubiquity of colloidal solutions in everyday industrial applications such as papermaking and coatings there is a need to be able to efficiently design and manufacture these substances. A related issue concerns the connection between many physiological diseases and heath defects and the stability and phase behavior of certain proteins. It is imperative to understand the physical mechanisms that cause proteins to change their normal solution characteristics. To design colloidal solutions for specific applications as well as to produce preventative medicines and therapies an intimate knowledge of the connection between particle interactions and overall physical properties of the solution is needed. To probe this issue four types of systems are examined. In each system solution conditions are altered affecting the nature and strength of the particle interactions. Our goal is to understand the physics behind the evolution of fluid properties that occurs because of changes in microscopic interactions. The method we employ in this pursuit is grand canonical transition matrix Monte Carlo. We examine an embedded point charge protein model of lysozyme in bulk, mixed with polymer, as well as in confinement. We find that in bulk the model is able to capture qualitatively experimental trends for changes in critical temperature and evolution of the fluid phase diagram with changing solution conditions such as salt concentration and pH. Quantitatively the model predicts a relatively narrow coexistence curve compared to experimental values. It is found that the osmotic second virial coefficient remains relatively constant over a broad range of solutions conditions suggesting a universal magnitude of attraction needed to induce phase separation. We examine a simple system consisting of hard sphere colloids with added Gaussian core polymers. Decreasing the size of the polymers relative to colloids as well as increasing the energetic repulsion between polymers upon overlap results in an overall stabilization of the mixture. Unlike bulk solutions containing molecules of the Carlsson et al. lysozyme model, the osmotic second virial coefficient at the critical point for model colloid-polymer mixtures is not constant but depends on polymer size and interaction. Increasing polymer size or decreasing polymer repulsion results in a larger negative value. Overall the model fails to capture the experimental behavior of polymer excluded volume interactions because its inability to describe the polymers capability of deformation around the colloid. We extend our analysis to a mixture containing the embedded charge model for lysozyme and Gaussian core polymers. Overall, the system exhibited a strong dependence on pH and salt concentration that qualitatively followed experimental trends. Increase of salt concentration or decrease in protein charge decreases the number of polymers needed to induce phase separation. This trend was not sensitive to the size of the polymer relative to the protein. Finally we examine the effect surface interactions have on the phase behavior for the lysozyme model as well as a simple square well model. Both systems exhibited a distinctly non-monotonic variation of its critical temperature as a function of fluid-wall interaction strength. A maximum occurs at an intermediate strength. We introduce two metrics that enable one to predict the location of this maximum. The first is related to the contact angle a fluid makes with the confining substrate while the second is based upon virial coefficient information. Because similar trends are exhibited in both systems we believe that the results should be general in nature.

This volume of Current Topics in Membranes focuses on Membrane Protein Crystallization, beginning with a review of past successes and general trends, then further discussing challenges of mebranes protein crystallization, cell free production of membrane proteins and novel lipids for membrane protein crystallization. This publication also includes tools to enchance membrane protein crystallization, technique advancements, and crystallization strategies used for photosystem I and its complexes, establishing Membrane Protein Crystallization as a needed, practical reference for researchers.

Providing the necessary basis for any developments of theoretical thermodynamic models, this book provides a complete collection of practical thermodynamic data for a variety of applications, including: basic and applied chemistry, chemical engineering, thermodynamic research, computational modeling, membrane science and technology, and environmental and green chemistry. The data -- which includes such developments as vapor-liquid and liquid-liquid equilibria, low-and high-pressure equilibrium data, enthalpic and volumetric data, and second virial coefficients -- is necessary when studying intermolecular interactions and gaining insights into the molecular nature of mixtures.

International Review of Cytology presents current advances and comprehensive reviews in cell biology-both plant and animal. Articles address structure and control of gene expression, nucleocytoplasmic interactions, control of cell development and differentiation, and cell transformation and growth. Authored by some of the foremost scientists in the field, each volume provides up-to-date information and directions for future research.This volume provides an overview of major cytoplasmic properties and events which including cytoarchitecture and the physical properties of cytoplasm, molecular compartmentation and gradients, channeling, sorting, and trafficking. It also addresses physicochemical events, both measured and anticipated, which attend solutions under conditions prevailing in cytoplasm: molecular crowding. It summarizes the current state of knowledge in the field and considers questions such as how molecules in cytoplasm interact.

Providing valuable insight on physical behavior of polymer solutions, intermolecular interactions, and the molecular nature of mixtures, each volume in this one-of-a-kind handbook brings together reliable, easy-to-use entries, references, tables, examples, and appendices on experimental data from hundreds of primary journal articles, dissertations,

This book covers the fundamentals of the rapidly growing field of biothermodynamics, showing how thermodynamics can best be applied to applications and processes in biochemical engineering. It describes the rigorous application of thermodynamics in biochemical engineering to rationalize bioprocess development and obviate a substantial fraction of t