PURIFICATION AND CHARACTERIZATION OF PROINSULIN ASPART ASPS.RWR

• Main Author: Pusvita Dewi, Sari
• Format: Final Project
• Language: Indonesia
• Online Access: https://digilib.itb.ac.id/gdl/view/64950
• Institution: Institut Teknologi Bandung

Insulin asprate is an insulin analog that is widely used for the treatment of diabetes mellitus in Indonesia. Generally, commercial insulin analogues are mostly produced by the host cells of Saccharomyces cerevisiae and Escherichia coli. Recently, researchers discovered the fact that proinsulin aspart can be produced in Pichia pastoris. Therefore, the aim of this study was to produce, purify and characterize proinsulin asprate. The experiment was started by producing proinsulin aspart at 30°C with methanol induction for 2 x 24 hours, purifying proinsulin aspart by cation exchange chromatography using CM-650M resin, then characterizing the protein by Western Blot method. The presence of a protein band measuring about 8.3 kDa on the SDS-PAGE electropherogram indicates that the proinsulin protein Aspart AspS.RWR has been successfully produced in Yeast Yeast Extract Peptone Dextrose (YPD), Buffered Glycerol Complex Medium (BMGY) and Buffered Metanol Complex Medium (BMMY). Meanwhile, the purification process for proinsulin Aspart AspS.RWR has been successfully carried out, with only one protein band found in the elution pathway resulting from SDS-PAGE. Western Blot analysis of the protein purification results showed that there were several bands that were sized according to the molecular weight of the proinsulin AspS.RWR. In essence, purification of proinsulin aspart by cation exchange chromatography, can separate proinsulin aspart protein from other proteins and proinsulin Aspart can be characterized by Western Blot.