Doctor of Philosophy

dissertationGlobally, it is estimated that more than 400 million people suffer from diabetes mellitus -- a chronic metabolic disease characterized by high blood glucose levels. Most individuals with diabetes depend on multiple administrations of different insulin analogues to maintain tight glycemic control and a healthy lifestyle. Currently, several insulin analogues are commercially available to treat diabetes and all are synthesized via recombinant expression. However, total chemical synthesis remains of great interest because it expands the therapeutic toolkit beyond the 20 natural amino acids. Chapter 1 provides an overview of diabetes with a focus on the biosynthetic and chemical synthesis as well as challenges faced by the synthesis of this peptide hormone. Chapters 2 and 3 describe several strategies to enzymatically and chemically synthesize new insulin analogues and derivatives for the treatment of diabetes. Specifically, Chapter 2 tackles current limitations in the methods used to synthesize insulin derivatives via the C-terminus of the B-chain. Furthermore, it introduces a new ligation method with high efficiency and functional group tolerance utilizing a bacterial transpeptidase, Staphylococcus aureus Sortase A (SrtA). This chapter demonstrates that SrtA can be used to generate new insulin derivatives in high yields (70-80%) and can be used as a facile method to ligate unique molecules to insulin. Chapter 3 addresses the challenges of the notoriously hydrophobic insulin A-chain and current methods to enhance its solubility, purification, and handling. Herein, a strategy is described that uses a solubility linker iv known as a "helping hand" that is coupled to the N-terminus of an insulin-based A-chain to enhance its solubility. Chapter 4 addresses the need for the development of a monomeric ultra-fast acting insulin (UFI) analogue to help improve glycemic control. Herein, we take inspiration from a UFI produced by the fish-hunting cone snail, Conus geographus, to synthesize venomous insulin molecules and analyze their structure-activity relationship to the human insulin receptor. Finally, Chapter 5 focuses on the conclusions and future directions for developing novel strategies that expand the insulin-based peptide therapeutics repertoire