Total Chemical Synthesis of SUMO-2-Lys63-linked diUbiquitin Hybrid Chains Assisted by Removable Solubilizing Tags

Along with the known posttranslational modification by ubiquitin, known as ubiquitination, there are many ubiquitin like modifiers such as the Small Ubiquitin Like Modifier (SUMO) proteins, which are know to regulate many important cellular processes. Like ubiquitination, SUMOlytion is also mediated by E1, E2, and E3 enzymes linked via an isopeptide bond to the C-terminal Gly of SUMO to a Lys residue from a target protein. Recently, the hybrid chains of SUMO-ubiquitin and specifically SUMO-2 linked to Lys63-di-ubiquitin were found to play major role in DNA repair. Despite some progress in understanding the role of the hybrid chains in DNA repair, there are various fundamental questions remained to be answered. To farther investigate the importance of hybrid SUMO-ubiquitin chains in DNA repair, homogenous material of the hybrid chains and their unique analogs are needed in workable quantities. For the first time and by applying advanced chemical strategies in protein synthesis we report the total chemical synthesis of four different SUMO-2-Lys63-linked di-ubiquitin hybrid chains. In this synthesis, the usefulness of removable solubilizing tags is demonstrated and new lessons were learned for future studies where peptide fragments are difficult to handle and purify. The availability of these chains open new opportunities in studying the role of these chains in DNA repair and other cellular processes, which we are currently pursuing.