Biotransformation of High Concentrations of Ginsenoside Substrate into Compound K by β-glycosidase from Sulfolobus solfataricus

Abstract

The rare ginsenoside Compound K (CK) is an attractive ingredient in traditional medicines, cosmetics, and the food industry because of its various biological activities. However, it does not exist in nature. The commonly used method for the production of CK is enzymatic conversion. In order to further improve the catalytic efficiency and increase the CK content, a thermostable β-glycosidase from Sulfolobus solfataricus was successfully expressed in Pichia pastoris and secreted into fermentation broth. The recombinant SS-bgly in the supernatant showed enzyme activity of 93.96 U/mg at 120 h when using pNPG as substrate. The biotransformation conditions were optimized at pH 6.0 and 80 °C, and its activity was significantly enhanced in the presence of 3 mM Li+. When the substrate concentration was 10 mg/mL, the recombinant SS-bgly completely converted the ginsenoside substrate to CK with a productivity of 507.06 μM/h. Moreover, the recombinant SS-bgly exhibited extraordinary tolerance against high substrate concentrations. When the ginsenoside substrate concentration was increased to 30 mg/mL, the conversion could still reach 82.5% with a productivity of 314.07 μM/h. Thus, the high temperature tolerance, resistance to a variety of metals, and strong substrate tolerance make the recombinant SS-bgly expressed in P. pastoris a potential candidate for the industrial production of the rare ginsenoside CK.

Optimization of Antioxidant Activity of Compounds Generated during Ginseng Extract Fermentation Supplemented with Lactobacillus

“…In the case where the content of Rg1 decreases while Rh 1 remains unchanged, it is possible that Rh4 or PPT, along with other unidentified and quantitatively undetermined saponins, have been produced ( Figure S1 ). The high concentration of saponin substrates can affect the hydrolytic ability of glycosidases towards saponin glycosidic bonds [ 48 ]. Thus, apart from Rg 5 , there was no significant changes in the content of detected rare ginsenosides before and after fermentation.…”

Compound K Production: Achievements and Perspectives

“…Consequently, there is a need to explore alternative microbial hosts for ginsenoside production. Recently, Pichia pastoris (P. pastoris) and Lactococcus lactis have been used as microbial hosts for β-glucosidase expression [72,73]. Both strains are generally regarded as safe (GRAS) microbes.…”

“…Therefore, these strains are a suitable expression system for heterologous protein production. In the case of lactic acid bacteria, L. lactis NZ9000, carrying β-glucosidase genes (BglPm and BglBX10) from Paenibacillus mucilaginosus and Flavobacterium johnsoniae, demonstrated the conversion of up to 70% of Rb1 to CK [73]. In another case, P. pastoris was used as a microbial host for the expression of a thermostable β-glucosidase from Sulfolobus solfataricus.…”

The Synthesis of Ginsenoside Compound K Using a Surface-Displayed β-Glycosidase Whole-Cell Catalyst

“…The mobile phase consisted of a gradient of water (A) and acetonitrile (B), as follows: 65% A, 0-10 min; 65% to 45% A, 10-12 min; 45% A, 12-35 min; 45% to 0% A, 35-40 min; 0% to 65% A, 40-45 min; and 65% A, 45-50 min. The flow rate was established at 1.5 mL/min [1].…”

“…Ginseng is a traditional herbaceous plant that can be used to maintain physical vitality and prolong life, so it is also known as the "king of herbs" [1]. The pharmacological activity of ginseng is mainly attributed to ginsenosides.…”