Pengaruh Variasi Konsentrasi Galaktosa dan Rafinosa Sebagai Sistem Induksi terhadap Produksi Enzim Endo -ß-1,4-D-Xilanase (XYNBTN63D) dalam Saccharomyses Cerevisiae BJ1824

Abstract

Endo-β-1,4-D-xylanase is an enzyme that hydrolyzes β-1,4-glycosidic bonds in xylan to produce xylooligosaccharides (XOS), which have potential as prebiotic compounds. This study aimed to determine the effects of galactose and raffinose concentration variations on the growth of Saccharomyces cerevisiae BJ1824 (pYHM1-xynBTN63D) and recombinant Endo-β-1,4-D-xylanase activity. Galactose was used as an inducer of the GAL1 promoter, while raffinose served as a non-repressing carbon source. Five media were evaluated, namely YPG, YMin 0%, YMin A (0.5% galactose and 1.5% raffinose), YMin B (1.5% galactose and 0.5% raffinose), and YMin 1% (1% galactose and 1% raffinose). Cell growth was monitored based on OD600, enzyme activity was determined using the DNS method, and hydrolysis products were characterized using thin-layer chromatography (TLC). The results showed that YMin B produced the highest biomass among the minimal media, with an OD600 of approximately 1.40 at the optimum phase. YMin B also showed the highest enzyme activity among the minimal media, reaching 6.460 U/mL, compared with 7.498 U/mL in YPG. In contrast, YMin 0% produced the lowest enzyme activity at 0.702 U/mL. TLC analysis showed that xylotriose was the dominant hydrolysis product in the YMin media containing galactose and raffinose, whereas YPG tended to produce products corresponding to xylobiose. Overall, YMin B containing 1.5% galactose and 0.5% raffinose was the best composition among the minimal media tested for supporting cell growth and recombinant Endo-β-1,4-D-xylanase activity. These findings indicate that galactose concentration plays an important role in induction, while raffinose supports cell growth as a non-repressing carbon source