SYNTHETIC PROTEIN DESIGN AND ENGINEERING OF A MONOCLONAL ANTIBODY

Abstract

Antibody engineering has enabled the directed modification of immunoglobulins to improve their biological action. This field of engineering has also enabled the construction of smaller, customized antibodies, termed antibody fragments. Engineered antibody fragments, are formed by fusing an antibody fragment and a multimerization domain using a peptide linker region. This thesis reports on the construction of a number of engineered antibody fragments, their expression in microbial host Escherichia coli, and purification by affinity chromatography. Furthermore, we investigated the effects of the linker on the structural properties and functionality of the engineered antibody fragment. Lastly, using engineered antibody fragments with a valency ranging from one to four, we observed an inverse exponential relationship of valency versus cytotoxicity, indicating that valency is the deciding factor in increasing cytotoxicity. In conclusion, the trimeric engineered antibody fragment was the optimum protein for targeting undifferentiated hESC with potential applications in regenerative medicine.