title: Metabolomic and transcriptomic analysis for rate-limiting metabolic steps in xylose utilization by recombinant Candida utilis.
abstract: We have reported that a recombinant Candida utilis strain expressing a Candida shehatae xylose reductase K275R/N277D, a C. shehatae xylitol dehydrogenase, and xylulokinase from Pichia stipitis produced ethanol from xylose, but its productivity was low. In the present study, metabolomic (CE-TOF MS) and transcriptomic (microarray) analyses were performed to characterize xylose metabolism by engineered C. utilis and to identify key genetic changes contributing to efficient xylose utilization. The metabolomic analysis revealed that the xylose-fermenting strain accumulated more pentose phosphate pathway intermediates, more NADH, and more glycolytic intermediates upstream of glyceraldehyde 3-phosphate than the wild-type. Transcriptomic analysis of the strain grown on xylose indicated a significant increase in expression of the genes encoding tricarboxylic acid cycle enzymes, respiratory enzymes, and enzymes involved in ethanol oxidation. To decrease the NADH/NAD(+) ratio and increase the ethanol yield of the fermentation of xylose, ADH1 encoding NADH-dependent alcohol dehydrogenase was overexpressed. The resulting strain exhibited a 17% increase in ethanol production and a 22% decrease in xylitol accumulation relative to control.
authors.affiliation: Central Laboratories for Key Technologies, KIRIN Company, Ltd, Yokohama, Kanagawa 236-0004, Japan. Hideyuki_Tamakawa@kirin.co.jp
authors.forename: Hideyuki
authors.initials: H
authors.lastname: Tamakawa
authors.forename: Yasuyuki
authors.initials: Y
authors.lastname: Tomita
authors.forename: Aki
authors.initials: A
authors.lastname: Yokoyama
authors.forename: Yuki
authors.initials: Y
authors.lastname: Konoeda
authors.forename: Shigehito
authors.initials: S
authors.lastname: Ikushima
authors.forename: Satoshi
authors.initials: S
authors.lastname: Yoshida
mesh_terms: D000426:Alcohol Dehydrogenase / Q000235:genetics; D002175:Candida / Q000166:cytology / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism*; D004274:DNA, Recombinant / Q000235:genetics*; D020869:Gene Expression Profiling; D005818:Genetic Engineering; D042541:Intracellular Space / Q000378:metabolism; D007700:Kinetics; D055432:Metabolomics; D014994:Xylose / Q000378:metabolism*
chemical_list: D004274:DNA, Recombinant; D014994:Xylose; D000426:Alcohol Dehydrogenase