Efficient production of ginsenoside Rh2 from xylose by remodeling metabolism in Saccharomyces cerevisiae

Ginsenoside Rh2, a promising candidate for cancer prevention and therapy, has applications in the fields of health products and pharmaceuticals. However, the current production of Rh2 through the extraction of Panax ginseng is not sustainable and cannot meet the increasing market demand. Here, we increased the Rh2 production in Saccharomyces cerevisiae by engineering the heterologous xylose metabolic pathway. The Candida tropicalis derived xylose reductase-xylitol dehydrogenase pathway was successfully constructed in in the previously engineered protopanaxadiol (PPD)-producing strain. A PPD high-yield strain with strong xylose utilization capacity was optimized to reduce the generation of byproduct xylitol with the expression of a soluble pyridine nucleotide transhydrogenase. The new yeast chassis produced 494.0 mg/L PPD in xylose medium, which could provide sufficient precursors for Rh2. The Rh2 production strain was then constructed by integrating the glycosyltransferases UGTPg45 overexpression module and further optimized through systematic enhancement of its mevalonic acid (MVA) pathway, resulting in a Rh2 titer of 209.5 mg/L in xylose medium. In addition, we confirmed that the engineered strain also exhibited good fermentation performance in the lignocellulose hydrolysate mimic glucose-xylose mixed medium, and we further elucidated the reasons of higher Rh2 production in xylose fermentation stage through transcriptomic and metabolic analysis. Taken all strategies together, the resulting strain produced 1.47 g/L Rh2 in a 5 L bioreactor with mixed sugar medium, making it possible for a green and sustainable biotechnology to produce ginsenoside Rh2 using xylose efficiently. This is the first report on the use of xylose for high-value chemical ginsenoside Rh2 production, which will pave the way for the future industrial production of ginsenoside Rh2 from a renewable carbon source such as lignocellulose.