EpCAM is a type I human transmembrane glycoprotein, expressed at low levels in epithelial cells and at high levels in a variety of stem cells. Additionally, high levels of expression have also been observed in various carcinomas. Due to the latter characteristic, EpCAM has been proposed as a potential target in treating certain types of cancer. Indeed, therapeutic antibodies have been developed. However, our understanding of EpCAM's involvement in cellular processes leaves much to be desired. After the structure of EpCAM's extracellular part had been experimentally determined, a hydrophobic pocket was observed on its surface. EpCAM's involvement in cellular processes is facilitated by regulated intramembrane proteolysis. We propose that the hydrophobic pocket could play a role in regulating the aforementioned process. Previously, we designed and prepared mutated forms of EpCAM with an obstructed pocket and qualitatively described changes in EpCAM's regulated intramembrane proteolysis. In this thesis, we quantified these changes and analyzed changes in the localization of mutant forms. We also expressed the former in an Sf9 insect cell line. We used the purified proteins to determine the ligand-binding capabilities of EpCAM's hydrophobic pocket, both in vitro and in silico. We observed the localization of mutant forms to unknown fibrous structures and quantified their changed susceptibility to regulated intramembrane proteolysis. Additionally, we provide proof of their correct folding and binding of ligands.