Part:BBa_K3003008

PelB-PenetraMax-SCI57-Own SCI57 Linker

Single Chain Insulin 57 (SCI57) is used for insulin expression in the E. coli with a linker peptide. SCI57 is insulin analog that demonstrates high activity and ultrastability. We took SCI-57 design from the study “Design of an active ultrastable single-chain insulin analog: synthesis, structure, and therapeutic implications” done by Hua QX et al. SCI-57 differs from other single chain insulin analogs by modification in isoelectric points of determined amino acids and it was aimed forisoelectric points to be near as insulin. Binding affinity of SCI57 is considerably higher than the wild-type insulin. Linker peptide provides SCI57 to be expressed in E. coli as a single chain form. Linker we have used between SCI57's b and a chain was SCI57’s own linker. This linker is produced for SCI57 for providing more bioactivity and affinity for SCI57 to recognized by the receptor, therefore it increases the affinity of SCI for binding to insulin receptor in a more efficient way. Progression of wild-type insulin is processed in a 3 stage. Production of Pre-Proinsulin, Proinsulin and Insulin. Proinsulin contains c peptide between alpha and beta chain as a linker. C peptide is cleaved by secretory vesicles found in Golgi apparatus and final form of insulin is secreted by Beta Cells in pancreas. Due to the inefficiency of E. coli to cleave this peptide, insulin analogs which are expressed in a single chain insulin form with peptide linker that provides this formation. PenetraMax is a cell penetrating peptide which included in this part. Cell penetrating peptide is used in this part for SCI57 to penetrate the intestinal epithelium for getting through the bloodstream. This peptide is taken from the study “One-month subchronic toxicity study of cell-penetrating peptides for insulin nasal delivery in rats” done by Khafagy et. al. PenetraMax facilitates the delivery of insulin from the intestine to the bloodstream. Tobacco Etch Virus (TEV) Linker part’s function is providing recognition sequence for TEV enzyme to cut the protein of interest (SCI57) for releasing and bridge between Ag43 Alpha Subunit to be displayed in the surface.