Enhancement of xylose uptake in 2-deoxyglucose tolerant mutant of Saccharomyces cerevisiae.
Chemical mutation of Saccharomyces cerevisiae using ethyl methane sulfonate was performed to enhance its ability of xylose uptake for ethanol production from lignocellulose under microaerobic condition. Among the appeared mutants, the mutant no. 2 (M2) strain screened using inhibitory effects of 2-deoxyglucose (DOG) showed more than 4-fold high ability in xylose uptake compared with the wild type strain, under the presence of glucose. The catabolite repression by glucose was sufficiently reduced in M2 strain due to its tolerance to the high concentration of DOG (0.5%, wt./vol.). Metabolomic analyses of various sugars in the cell revealed that some of xylose was reduced to xylitol in M2 cell, providing the concentration gradient of xylose and more uptake of xylose. Xylulose-5-phosphate was significantly detected in the crude cell extract from M2 strain, indicating higher metabolic activity in pentose phosphate pathway. This was also confirmed by in vitro analyses of key enzymes involved in glucose and xylose metabolism, such as hexokinase, glucose-6-phosphate dehydrogenase and xylose reductase. Glucose uptake was moderately suppressed in the presence of trehalose-6-phosphate inhibiting the activation of hexokinase, resulting in more uptake of xylose through hexose transport system. To our knowledge, this study is the first report verifying that the mutation technique successfully enhances the xylose uptake by S. cerevisiae, particularly under the presence of glucose.
Asian Center for Environmental Research, Meisei University, Tokyo, Japan. pri@hino.meisei-u.ac.jp
Prihardi
P
Kahar
Kazuo
K
Taku
Shuzo
S
Tanaka
D000449:Aldehyde Reductase / Q000235:genetics / Q000378:metabolism; D057465:Catabolite Repression; D003847:Deoxyglucose / Q000378:metabolism*; D000431:Ethanol / Q000378:metabolism; D005285:Fermentation; D005947:Glucose / Q000378:metabolism; D005954:Glucosephosphate Dehydrogenase / Q000378:metabolism; D006593:Hexokinase / Q000378:metabolism; D055432:Metabolomics; D016296:Mutagenesis; D009154:Mutation; D010428:Pentosephosphates / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000235:genetics / Q000378:metabolism*; D013403:Sugar Phosphates / Q000378:metabolism; D014199:Trehalose / Q000031:analogs & derivatives / Q000378:metabolism; D014993:Xylitol / Q000378:metabolism; D014994:Xylose / Q000378:metabolism*
D010428:Pentosephosphates; D013403:Sugar Phosphates; D000431:Ethanol; C082722:trehalose-6-phosphate; C031625:xylulose-5-phosphate; D003847:Deoxyglucose; D014994:Xylose; D014199:Trehalose; D000449:Aldehyde Reductase; D005954:Glucosephosphate Dehydrogenase; D006593:Hexokinase; D005947:Glucose; D014993:Xylitol