Date of Award
4-21-2009
Degree Type
Thesis
Degree Name
Master of Science (MS)
Department
Chemistry
First Advisor
Jenny J. Yang - Chair
Second Advisor
Zhi-Ren Liu
Third Advisor
Gangli Wang
Fourth Advisor
Giovanni Gadda
Abstract
Previous studies in Dr. Yang’s laboratory have established a grafting, design, and subdomain approach in order to investigate the properties behind Ca2+-binding sites located in Ca2+-binding proteins by employing engineered proteins. These approaches have not only enabled us to isolate Ca2+-binding sites and obtain their Ca2+-binding affinities, but also to investigate conformational changes and cooperativity effects upon Ca2+ binding. The focus of my thesis pertains to optimizing the expression and purification of engineered proteins with tailored functions. Proteins were expressed in E. coli using different cell strains, vectors, temperatures, and inducer concentrations. After rigorous expression optimization procedures, proteins were further purified using chromatographic and/or refolding techniques. Expression and purification optimization of proteins is essential for further analyses, since the techniques used for these studies require high protein concentrations and purity. Evaluated proteins had yields between 5-70 mg/L and purities of 80-90% as confirmed by SDS-PAGE electrophoresis.
DOI
https://doi.org/10.57709/1059241
Recommended Citation
Castiblanco, Adriana P., "Expression and Purification of Engineered Calcium Binding Proteins." Thesis, Georgia State University, 2009.
doi: https://doi.org/10.57709/1059241