Figure captions
Fig. 1A. Biosynthetic pathway of β-carotene from glucose and xylose in engineered S. cerevisiae . A heterogolous xylose assimilation pathway containing xylose reductase (XR), xylitol dehydrogenase (XDH) and xylulokinase (XK) is connected with lower glycolytic pathway by pentose phosphate pathway (PPP). Pyruvate is produced from glucose and xylose and converted into cytosolic acetyl-CoA. Yeast synthesizes farnesyl pyrophosphate from cytosolic acetyl-CoA through the mevalonate pathway (MVA), as the common precursor for the biosynthesis of ergosterol and heterogolous β-carotene. HMG-CoA reductase (HMGR) is a key rate limiting enzyme in MVA. Cytosolic acetyl-CoA is also the precursor for yeast lipids synthesis. G6P, glucose-6-phosphate; F6P, fructose-6-phosphate; GAP, glyceraldegyde-3-phosphate; X5P, xyllulose-5-phosphate; IPP, isopentenyl pyrophosphate; DMAPP, dimethylallyl pyrophosphate; GPP, geranyl pyrophosphate; FPP, farnesyl pyrophosphate; GGPP, geranylgeranyl pyrophosphate. Fig. 1B. Pictures of glucose and xylose cultures of SR8B strain at 30 hour. Fig. 1C. Transcriptional levels of the genes related to the production of β-carotene in engineered S. cerevisiae SR8B on glucose and xylose. Samples were taken at exponetial phase from each condition for RNA extraction and expression level analysis. Fold changes were calculated by dividing genes expression levels on xylose by those on glucose. Three biological replicates and three technical replicates were performed for each gene, and the error bars represented standard deviations.
Fig. 2. The overlaid HPLC chromatograms of carotenoids extracted from the engineered S. cerevisiae SR8B cultured on glucose and xylose condition. Cells were cultured in a defined medium containing either 40 g/L glucose or 40 g/L xylose from initial OD600 1. Same amount of cells were harvested at 90 hour from each condition to extract the carotenoids for HPLC chromotography.
Fig. 3. Batch fermentation profiles of the engineered S. cerevisiae SR8B on glucose (A) and xylose (B) conditions and the corresponding β-carotene production patterns on glucose (C) and xylose (D). Data are presented as mean values and standard deviations of three independent biological replicates.
Fig. 4 . Comparison of β-carotene production by the engineered SR8B and SR8BH strains through glucose and xylose utilization. Cells were harvested at the end of fermentation for β-carotene extraction and quantification. Data are presented as mean values and standard deviations of three independent biological replicates
Fig. 5A . Ergosterol production by the engineered SR8B strain through glucose and xylose utilization. Cells were harvested at the end of fermentation for ergosterol extractiona and quantification. Data are presented as mean values and standard deviations of three independent biological replicates. Fig. 5B. Lipid bodies visualization of the engineered SR8B cells cultured in glucose and xylose condition. Cells were harvested at exponential phase and stained with Nile Red fluorescent dye. The stained cells were then viewd under a confocal microscope with 63× oil immersion objective at 633 nm.
Fig. 6 Xylose feb-batch fermentation of the SR8B strain in a 3-litter bioreactor. The bioreactor picture on the right was taken at 80 h.