Increased expression of the oxidative pentose phosphate pathway and gluconeogenesis in anaerobically growing xylose-utilizing Saccharomyces cerevisiae.
BACKGROUND
Fermentation of xylose to ethanol has been achieved in S. cerevisiae by genetic engineering. Xylose utilization is however slow compared to glucose, and during anaerobic conditions addition of glucose has been necessary for cellular growth. In the current study, the xylose-utilizing strain TMB 3415 was employed to investigate differences between anaerobic utilization of glucose and xylose. This strain carried a xylose reductase (XYL1 K270R) engineered for increased NADH utilization and was capable of sustained anaerobic growth on xylose as sole carbon source. Metabolic and transcriptional characterization could thus for the first time be performed without addition of a co-substrate or oxygen.


RESULTS
Analysis of metabolic fluxes showed that although the specific ethanol productivity was an order of magnitude lower on xylose than on glucose, product yields were similar for the two substrates. In addition, transcription analysis identified clear regulatory differences between glucose and xylose. Respiro-fermentative metabolism on glucose during aerobic conditions caused repression of cellular respiration, while metabolism on xylose under the same conditions was fully respiratory. During anaerobic conditions, xylose repressed respiratory pathways, although notably more weakly than glucose. It was also observed that anaerobic xylose growth caused up-regulation of the oxidative pentose phosphate pathway and gluconeogenesis, which may be driven by an increased demand for NADPH during anaerobic xylose catabolism.


CONCLUSIONS
Co-factor imbalance in the initial two steps of xylose utilization may reduce ethanol productivity by increasing the need for NADP+ reduction and consequently increase reverse flux in glycolysis.
Department of Applied Microbiology, Lund University, PO Box 124, SE-221 00 Lund, Sweden. david.runquist@tmb.lth.se
David
D
Runquist
Bärbel
B
Hahn-Hägerdal
Maurizio
M
Bettiga
D000332:Aerobiosis; D000449:Aldehyde Reductase / Q000235:genetics / Q000378:metabolism; D000693:Anaerobiosis; D000431:Ethanol / Q000378:metabolism; D005285:Fermentation; D005818:Genetic Engineering / Q000379:methods; D005943:Gluconeogenesis / Q000235:genetics*; D005947:Glucose / Q000378:metabolism; D009249:NADP / Q000378:metabolism; D010427:Pentose Phosphate Pathway / Q000235:genetics*; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000378:metabolism; D015854:Up-Regulation; D014994:Xylose / Q000378:metabolism*
D000431:Ethanol; D009249:NADP; D014994:Xylose; D000449:Aldehyde Reductase; D005947:Glucose