Effects of Charge Distribution on Protein Solution Properties

In this study, the electrostatic properties of a series of proteins are characterized experimentally and related to theoretical predictions. The strength of protein interactions is characterized as a function of solute concentration, pH, and ionic strength. The strength of interaction and the solubility of lysozyme are characterized in the presence of three common cryoprotectants. The dramatic decrease in attractions and increase in solubility due to these agents highlights how poorly understood the actions of solutes on protein interactions are and that in design protein separation technologies measurements in the exact buffer conditions of interest are required. The net charge on lysozyme and a closely related series of BPTI mutants is related to the electrophoretic mobility and conductivity increment measurements. Charge determined from electrophoresis agreed well with theoretical predictions in some cases and showed strong evidence of ion binding in others. Interpretation of the results of conductivity experiments was complicated by the influence of counterions in the protein solutions. Despite this, the agreement theory and experiment is remarkable good. The charge distribution of these proteins was explored using dielectric spectroscopy. Using theory typically applied to proteins yields results that differ significantly from theoretical predictions in some cases, particularly where the protein net charge is high. These results provide motivation for the development of theory that will include ion relaxations and the protein dipole moment, both of which are expected to contribute significantly under the measurement conditions. Second virial coefficients were measured and compared to predictions based on models of particle interactions including dipolar interactions. Using the charge determined by electrophoresis allows accurate prediction of the second virial coefficient in cases where ion binding is significant. The dipole moment can be significant in the interactions of proteins at low ionic strengths, where the range of electrostatic interactions is large. Including the dipole moment allows better qualitative prediction of the second virial coefficient at low ionic strength, particularly when the net charge of the protein is reduced.

Name thesis:degree_name

Ph.D.

Level thesis:degree_level

Dissertation

Discipline thesis:degree_discipline

Chemical Engineering

Grantor

University of Illinois at Urbana-Champaign

Year dc:date

2015

Author dc:creator

• Farnum, Michael Alan

Contributors dc:contributor

• Zukoski, Charles F.

• Chemistry, Physical

Language dc:language

eng

Identifier

(MiAaPQ)AAI9955611

OAI identifier oai:identifier

oai:www.ideals.illinois.edu:2142/82477