KONSTRUKSI DAN KARAKTERISASI MUTANMUTAN PROTEIN DISULFIDA ISOMERASE RAGI YANG TIDAK MENGANDUNG DOMAIN B

• Main Author: ZUSFAHAIR
• Format: Theses
• Language: Indonesia
• Subjects: Kimia
• Online Access: https://digilib.itb.ac.id/gdl/view/2301
• Institution: Institut Teknologi Bandung

Protein disulfide isomerase (PDI) is a multi-functional enzyme involved in catalyzing reduction, oxidation, and isomerization reactions of disulfide bonds. The aim of the research was to indentify domains responsible in the substrate binding in yeast PDI. Human Protein Disulfide Isomerase consists of five domain a, b, b\', a\' and c. Domain b and b\' has been proposed to function as the most essential domain for substrate binding. Three mutans that lack of putative substrate binding site had been constructed by inverted PCR method. The constructed mutans were designated as pdil (A109-166), pdil(A197-250), pdil(A109-250). Sodium Dodecyl sulfate - polyacrylamide gel electrophoresis (SDS-PAGE) analysis showed that the three mutants were not overexpressed. However, western blot analysis indicated that the three mutants were expressed in E. coli BL 21(DE3). The three mutans had only 11% of insulin reductase activity compared to that of the wild type. Further analysis of the pdil (A109-166) suggested that the low level expression of pdi mutant might not be caused of T7 promotor inactivation.