Enhanced Ergosterol Production By Recombinant S.cerevisiae Harboring Vitreoscilla Hemoglobin Gene (vgb)

In order to increase the ergosterol production, new methods have been carried out in this study that expressing Vitreoscilla hemoglobin gene (vgb), which has the function of trapping extra cellular oxygen to inside, in S. cerevisiae to increase the oxygen and energy supply. Recombinant plasmid pVgb-kanMX4 was constructed and transformed into 5. cerevisiae 1190 and recombinant plasmid pVgb-EXl transformed into S. cerevisiae.YSR 6.126-1 B. Furthermore, vgb was expressed in both tranformed cells. With sufficient oxygen supply, the ergosterol contents of recombinant S. cerevisiae 1190 (pVgb-KanMX4) and wild type S. cerevisiae 1190 grown in shake flasks were 1.14% and 0.61%, respectively. Under oxygen limitation condition, ergosterol contents in recombinant S. cerevisiae 1190 (pVgb-KanMX4) and wild type 5. cerevisiae 1190 were reduced to 0.28% and 0.41%, respectively. The content of ergosterol in S. cerevisiae.YSH 6.126-1 B was less, but when cultured under oxygen-rich condition and oxygen-limited condition its ergosterol contents were 200% and 280% that of the wild type, respectively. In a 30 hours fermentation study conducted in a 5 liter fermentor, 15.1 g/L Cell Dry Weight (CDW) containing 1.38% ergosterol was obtained from the culture of recombinant S. cerevisiae 1190 (pVgb-KanMX4); Only 14.8 g/L CDW containing 0.9% ergosterol was produced by wild type S. cerevisiae 1190. These results demonstrated that vgb played an important role in enhancing ergosterol production of S. cerevisiae.Under some experimental about the relationship of ceramides and oxygen hypothesis, studying the effect of vgb on the yield of ceramides was conducted in S. cerevisiae. Some imporvement was made in TLC analytical method of ceramides, and confirming the developing solvent is Chloroform:methanol:acetic acid (19:1:0.1).