Total Chemical Synthesis of Palmitoyl-Conjugated Insulin

Commercially available insulins are manufactured by recombinant methods for the treatment of diabetes. Long-acting insulin drugs (e.g., detemir and degludec) are obtained by fatty acid conjugation at LysB29 epsilon-amine of insulin via acid-amide coupling. There are three amine groups in insulin, and they all react with fatty acids in alkaline conditions. Due to the lack of selectivity, such conjugation reactions produce non-desired byproducts. We designed and chemically synthesized a novel thiol-insulin scaffold (CysB29-insulin II), by replacing the LysB29 residue in insulin with the CysB29 residue. Then, we conjugated a fatty acid moiety (palmitic acid, C16) to CysB29-insulin II by a highly efficient and selective thiol-maleimide conjugation reaction. We obtained the target peptide (palmitoyl-insulin) rapidly within 5 min without significant byproducts. The palmitoyl-insulin is shown to be structurally similar to insulin and biologically active both in vitro and in vivo. Importantly, unlike native insulin, palmitoyl-insulin is slow and long-acting.

Automated summary

Commercially available insulins for diabetes treatment are produced using recombinant methods. We developed a novel thiol-insulin scaffold (CysB29-insulin II) by substituting the LysB29 residue in insulin with CysB29 residue, and conjugated it with a fatty acid moiety (palmitic acid, C16) using a highly efficient and selective thiol-maleimide conjugation reaction. The resulting palmitoyl-insulin showed structural similarity to insulin, maintained biological activity, and exhibited slow and long-acting effects unlike native insulin.