• Journal of Microbiology and Biotechnology
• /
• v.10 no.4
• /
• pp.564-567
• /
• 2000

A recombinant Saccharomyces cerevisiae, transformed with the genes encoding xylose reductase (XYL1) and xylitol dehydrogenase (XYL2) orginated from Pichia stipitis CBS 5776, was developed to directly convert xylose to ethanol. A fed-batch fermentation with the recombinant yeast produced 8.7 g ethanol/l with a yield of 0.13 g ethanol/g xylose consumed.

• Journal of Life Science
• /
• v.27 no.12
• /
• pp.1403-1409
• /
• 2017

In this study, the xylitol dehydrogenase (XYL2) gene was expressed in Saccharomyces cerevisiae as a host cell for ease of use in the degradation of lignocellulosic biomass (xylose). To select suitable expression systems for the S.XYL2 gene from S. cerevisiae and the P.XYL2 gene from Pichia stipitis, $pGMF{\alpha}-S.XYL2$, $pGMF{\alpha}-P.XYL2$, $pAMF{\alpha}-S.XYL2$ and $pAMF{\alpha}-P.XYL2$ plasmids with the GAL10 promoter and ADH1 promoter, respectively, were constructed. The mating factor ${\alpha}$ ($MF{\alpha}$) signal sequence was also connected to each promoter to allow secretion. Each plasmid was transformed into S. cerevisiae $SEY2102{\Delta}trp1$ strain and the xylitol dehydrogenase activity was investigated. The GAL10 promoter proved more suitable than the ADH1 promoter for expression of the XYL2 gene, and the xylitol dehydrogenase activity from P. stipitis was twice that from S. cerevisiae. The xylitol dehydrogenase showed $NAD^+$-dependent activity and about 77% of the recombinant xylitol dehydrogenase was secreted into the periplasmic space of the $SEY2102{\Delta}trp1/pGMF{\alpha}-P.XYL2$ strain. The xylitol dehydrogenase activity was increased by up to 41% when a glucose/xylose mixture was supplied as a carbon source, rather than glucose alone. The expression system and culture conditions optimized in this study resulted in large amounts of xylitol dehydrogenase using S. cerevisiae as the host strain, indicating the potential of this expression system for use in bioethanol production and industrial applications.

• Korean Journal of Food Science and Technology
• /
• v.28 no.6
• /
• pp.1151-1156
• /
• 1996

Effects of xylose and glucose on the xylitol production were investigated with Candida parapsilosis KFCC 10875. With increasing the ratio of glucose to xylose, xylitol production decreased but ethanol and glycerol production increased. The maximum concentrations of ethanol and glycerol were 21.5 g/l and 3.6 g/l, respectively, in a medium consisting of 10 g/l xylose and 40 g/l glucose. No xylitol was formed in the glucose medium without xylose since xylitol could not be produced from glucose alone. The inhibitory effect of ethanol, a major by-product, on xylitol production was also studied. As the added ethanol concentration was increased, xylitol production decreased. When cells were inoculated in a xylose medium after removing the by-product (ethanol), xylitol production was not inhibited. The concentrated cells grown on xylose or glucose were inoculated in a fermentor containing the xylose medium. The total activities $(specific{\;}activities{\times}\;cell\;concentration)$ of xylose reductase and xylitol dehydrogenase in concentrated cells grown on glucose were the same as those in a normal fermentation; the specific activities of the above enzymes in the cells grown on xylose were the same as those in a normal fermentation. It indicates that the xylitol productivity of concentrated cells grown on xylose could be increased with increasing the cell concentration. By using concentrated cells of 20 g/l grown on xylose, the final xylitol concentration of 40 g/l was obtained for 18 h fermentation from 50 g/l xylose.

• Journal of Microbiology and Biotechnology
• /
• v.11 no.4
• /
• pp.564-569
• /
• 2001

This study was carried out in order to investigate the characteristics of xylitol fermentation by a xylitol dehydrogenase defective mutant PXM-4 of P stipitis CBS 5776 and to determime optimum conditions for the high yield ofxylitol production from xylose. Gluconic acid was selected as a co substrate for the xylitol fermentation, since gluconic acid neither blocked xylose transport nor repressed xylose reductase expression. An increase of gluconic acid concentration reduced the rates of xylitol production and cell growth by decreasing medium pH, and the optimal concentration of gluconic acid was determined to be 20 gll with approximately 100% xylitol conversion yield. A fed-batch cell culture resulted in a 44.8 g/l xylitol concentration with 100% yield, based on the amount of xylose consumed.

• Journal of Microbiology and Biotechnology
• /
• v.13 no.4
• /
• pp.501-508
• /
• 2003

Artificial coupling of one enzyme with another can provide an efficient means for the production of industrially important chemicals. Xylose reductase has been recently discovered to be useful in the reductive production of xylitol. However, a limitation of its in vitro or in vivo use is the regeneration of the cofactor NAD(P)H in the enzyme activity. In the present study, an efficient process for the production of xylitol from D-xylose was established by coupling two enzymes. A NADH-dependent xylose reductase (XR) from Pichia stipitis catalyzed the reduction of xylose with a stoichiometric consumption of NADH, and the resulting cofactor $NAD^+$ was continuously re-reduced by formate dehydrogenase (FDH) for regeneration. Using simple kinetic analyses as tools for process optimization, suitable conditions for the performance and yield of the coupled reaction were established. The optimal reaction temperature and pH were determined to be about $30^{\circ}C$ and 7.0, respectively. Formate, as a substrate of FDH, affected the yield and cofactor regeneration, and was, therefore, adjusted to a concentration of 20 mM. When the total activity of FDH was about 1.8-fold higher than that of XR, the performance was better than that by any other activity ratios. As expected, there were no distinct differences in the conversion yields of reactions, when supplied with the oxidized form $NAD^+$ instead of the reduced form NADH, as a starting cofactor for regeneration. Under these conditions, a complete conversion (>99%) could be readily obtained from a small-scale batch reaction.

• Microbiology and Biotechnology Letters
• /
• v.25 no.5
• /
• pp.495-500
• /
• 1997

Xylitol production from xylose and glucose was investigated using Candida tropicalis KFCC-10960. As glucose concentration in xylose medium was increased, ethanol production increased. However, xylitol production was maximum at glucose concentration of 10 g/l. The concentrated cells grown on xylose or glucose were inoculated in xylose medium. The specific activities of xylose reductase and xylitol dehydrogenase, and xylitol production in concentrated cells grown on glucose were the same as those in concentrated cells grown on xylose. The results suggested that cells grown on glucose had the same xylitol producing activity as those grown on xylose. By feeding glucose in xylose medium, cell growth was achieved from glucose and xylitol production was obtained from xylose. By using this technique, a final xylitol concentration of 261 g/l was achieved from 300 g/l xylose in 41 hours which corresponded to a xylitol yield from xylose of 87% and a xylitol productivity of 6.37 g/1-h.

• Current Research on Agriculture and Life Sciences
• /
• v.5
• /
• pp.68-74
• /
• 1987

The growth of A calcoaceticus C10 in CL medium was not increased by the addition of 0.5% E-capralactone or succinate, but increased by 0.2% adipate, xylose or even glucose which was not metabolized as a carbon source. The addition of 0.2% glucose after culture in CL medium for 6 hours increased the growth of A. colcoaceticus C10 twice as much as that in CL medium after culture for further 10 hours. Biosynthesis of cyclohexanol dehydrogenase in A. calcoaceticus C10 was not repressed by ${\varepsilon}$-caprolactone, succinate, xylose and glucose, but repressed by adipate which is endproduct of cyclohexanol metabolism. The induction of dehydrogenase by cyc1ohexanol in CL medium was not repressed completely by 0.1% adipate, but repressed almost completely by 0.2% adipate in A. calcoaceticus C10.

• KSBB Journal
• /
• v.15 no.2
• /
• pp.113-119
• /
• 2000

In order to produce xylitol with high yield, experiments were carried out to develope xylitol dehydrogenase (XDH) defective m mutant from Pichia stipitis and to investigate the xylit이 fermentation characteristics of mutant strain. After treatment of P s stipitis with EMS, mutant PXM-4 was selected based on the XDH activity and xylitol production capability. Among the tested c cosubstrates, galactose was selected as an adequate cosubstrate on xyl뻐I production of mutant PXM-4. With the increase of galactose concentration, xylitol production was decreased because the transport of xylose into cell was inhibited by g galactose. The optimal concentration of galactose for the production of xylitol using 20 g/L xylose was 20 g/L. Under this c condition, maximum concentration of xylitol and yield were 14.4 g/L and 97%, respectively. In order to prevent the inhibitory e effect of xylose transport by galactose, galactose was fed with low concentration and the concentration of xylitol produced w was increased up to 25 g/L. In the fermentation of corn cob hydrolyzate by mutant PXM-4, xylose was completely converted t to xylit이 with a 100% yield in 4 days culture.

• Korean Journal of Microbiology
• /
• v.37 no.2
• /
• pp.170-175
• /
• 2001

In order to produce xylitol with high yield, experiments were carried out to develope xylitol dehydrogenase(XDH) defective mutant from P. stipitis and to investigate the xylitol fermentation characteristics of mutant strain. After treatment of P. stipitis with EMS, mutant PXM-4 was selected based on te XDH activity and xylitol production capability. Among the tested cosubstrates, galactose was selected as an adequate cosub-strate on xylitol production of mutant PXM-4. But with the increase in the concentration of galactose in the medium, xylitol production was decreased because the transport of xylose into cell was inhibited by galactose. The optimal concentration of galactose for the production of xylitol using 20 g/ι xylose was 20 g/ι Under this condition, maximum concentration of xylitol and yield were 14.4 g/ι and 97%, respectively. In order to prevent the inhibitory effect of xylose transport by galactose, galactose was fed with low concentration and the concentration of xylitol produced was increased up to 25 g/ι.

• KSBB Journal
• /
• v.13 no.1
• /
• pp.6-12
• /
• 1998

In order to produce xylitol from hemicellulose hydrolysate which is widely used as a substrate, the development of strain such as catabolite derepressed mutant is required. After treatment of Candida sp. with EMS, GM-17 and PM-34 as catabolite derepressed mutant were isolated from Candida guilliermondii and Candida parapsilosis, respectively. Mutant GM-17 and PM-34 simultaneously assimilated xylose and glucose during the fermentation. The specific xylose reductase and xylitol dehydrogenase activities of mutant strains were also higher than those of wild strains in glucose medium and mixed medium of glucose and xylose. The xylitol productivity and yield of mutant GM-17 and PM-34 were improved as compared to the wild types in the mixed medium. The xylitol productivity and yield of mutant GM-17 were 0.09 g/L·hr and 0.56 g-xylitol/g-xylose, and those of mutant PM-34 were 0.21 g/L·hr and 0.52 g-xylitol/g-xylose in the mixed medium, respectively.