Omniligase-1-Mediated Phage-Peptide Library Modification and Insulin Engineering

Abstract

Chemical and enzymatic modifications of peptide-displayed libraries have been successfully employed to expand the phage display library. However, the requirement of specific epitopes and scaffolds has limited the scope of protein engineering using phage display. In this study, we present a novel approach utilizing omniligase-1-mediated selective and specific ligation on the phage pIII protein, offering a high conversion rate and compatibility with commercially available phage libraries. We applied this method to perform high-throughput engineering of insulin analogues with randomized B chain C-terminal regions. Insulin analogues with different B chain C-terminal segments were selected and exhibited biological activity equivalent to that of human insulin. Molecular dynamics studies of insulin analogues revealed a novel interaction between the insulin B27 residue and insulin receptor L1 domain. In summary, our findings highlight the potential of omniligase-1-mediated phage display in the development and screening of disulfide-rich peptides and proteins. This approach holds promise for the creation of novel insulin analogues with enhanced therapeutic properties and exhibits potential for the development of other therapeutic compounds.

Unraveling Light-Activated Insulin Action in Regulating Blood Glucose: New Photoactivatable Insight as a Novel Modality in Diabetes Management

“…Insulin is a protein secreted by the beta cells of the pancreas as a response to glucose ingestion to return the plasma glucose concentration to normal within 2 h [ 15 , 19 , 35 ]. Insulin is a relatively small protein with two peptide chains linked by disulfide bonds.…”

“…It stimulates vasodilation in arterial smooth muscle, enhancing blood flow, and the delivery of insulin and glucose to muscle cells [ 15 , 16 , 17 ]. Insulin is a relatively small protein with two peptide chains linked by disulfide bonds [ 18 , 19 ].…”

“…It stimulates vasodilation in arterial smooth muscle, enhancing blood flow, and the delivery of insulin and glucose to muscle cells [15][16][17]. Insulin is a relatively small protein with two peptide chains linked by disulfide bonds [18,19]. In T2D, insulin resistance occurs when cells, particularly the insulin receptors in the muscle, fat, and liver cells, do not respond adequately to insulin [20,21].…”