| Ubiquitination is a post translational protein modification which regulates a myriad of eukaryotic cellular functions. Ubiquitination targets the tagged protein for destruction by the 26S proteasome, or modulates protein activities, protein-protein interactions, or subcellular localization. The ubiquitination pathway involves three components, ubiquitin activating enzyme (E1), ubiquitin conjugating enzymes (E2s) and ubiquitin ligases (E3s), which are responsible for activation, covalent attachment and substrate recognition respectively. An understanding of the components involved in protein ubiquitination is essential to understand how specificity and regulation are conferred upon this pathway. The research reported in this thesis identifies interactions between E2s in the model system Caenorhabditis elegans. E2s, which form the key enzymes in ubiquitination or ubiquitin like modifications of the proteins, exist as ubiqtuin conjugating enzymes (UBCs) and ubiquitin conjugating enzyme variants (UEVs). The UEV proteins lack the critical cysteine residue necessary for conjugation which is present in the UBCs. In this study, the yeast two hybrid assay was employed to identify the interactions between the E2 proteins. It is interesting that the interactions identified involve the UEV proteins with UBC proteins. The interactions between the UEV protein, UEV-1, and the UBC proteins UBC-7, UBC-13, UBC-18 and UBC-25 were identified in the yeast two hybrid screen. Another UEV protein, UEV-2, is identified to interact with the UBC proteins, UBC-1 and UBC-6. The interaction between UEV-2 and UBC-1 is confirmed by pull down assay which may reveal a new role for UEV-2. The interactions observed in this research were novel and may shed light on the role of E2 dimerization in the ubiquitination pathway. |
