Xylitol production using recombinant Saccharomyces cerevisiae containing multiple xylose reductase genes at chromosomal delta-sequences.
Xylitol production from xylose was studied using recombinant Saccharomyces cerevisiae 2805 containing xylose reductase genes (XYL1) of Pichia stipitis at chromosomal delta-sequences. S. cerevisiae 2805-39-40, which contains about 40 copies of the XYL1 gene on the chromosome, was obtained by a sequential transformation using a dominant selection marker neor and an auxotrophic marker URA3. The multiple XYL1 genes were stably maintained on the chromosome even after 21 and 10 days in the non-selective sequential batch and chemostat cultures, respectively, whereas S. cerevisiae 2805:pVTXR, which harbors the episomal plasmid pVTXR having the XYL1 gene, showed mitotic plasmid instability and more than 95% of the cells lost the plasmid under the same culture conditions. In the first batch (3 days) of the sequential batch culture, volumetric xylitol productivity was 0.18 g l-1 h-1 for S. cerevisiae 2805-39-40, as compared to 0.21 g l-1 h-1 for S. cerevisiae 2805:pVTXR. However, the xylitol productivity of the latter started to decrease rapidly in the third batch and dropped to 0.04 g l-1 h-1 in the seventh batch, whereas the former maintained the stable xylitol productivity at 0.18 g l-1 h-1 through the entire sequential batch culture. The xylitol production level in the chemostat culture was about 8 g l-1 for S. cerevisiae 2805-39-40, as compared to 2.0 g l-1 for S. cerevisiae 2805:pVTXR after 10 days of cultures even though the xylitol production level of the latter was higher than that of the former for the first 5 days. The results of this experiment indicate that S. cerevisiae containing the multiple XYL1 genes on the chromosome is much more efficient for the xylitol production in the long-term non-selective culture than S. cerevisiae harboring the episomal plasmid containing the XYL1 gene.
Kim
Y S
YS
Department of Biological Sciences, Korea Advanced Institute of Science and Technology, Taejon, South Korea.
Kim
S Y
SY
Kim
J H
JH
Kim
S C
SC
D000449:Aldehyde Reductase / Q000235:genetics* / Q000378:metabolism; D015139:Blotting, Southern; D018628:Gene Dosage; D005800:Genes, Fungal; D005810:Multigene Family; D020161:Physical Chromosome Mapping; D010843:Pichia / Q000235:genetics; D010957:Plasmids / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism; D014170:Transformation, Genetic; D014993:Xylitol / Q000378:metabolism*