Isolation and characterization of the Pichia stipitis xylitol dehydrogenase gene, XYL2, and construction of a xylose-utilizing Saccharomyces cerevisiae transformant.
A P. stipitis cDNA library in lambda gt11 was screened using antisera against P. stipitis xylose reductase and xylitol dehydrogenase, respectively. The resulting cDNA clones served as probes for screening a P. stipitis genomic library. The genomic XYL2 gene was isolated and the nucleotide sequence of the 1089 bp structural gene, and of adjacent non-coding regions, was determined. The XYL2 open-reading frame codes for a protein of 363 amino acids with a predicted molecular mass of 38.5 kDa. The XYL2 gene is actively expressed in S. cerevisiae transformants. S. cerevisiae cells transformed with a plasmid, pRD1, containing both the xylose reductase gene (XYL1) and the xylitol dehydrogenase gene (XYL2), were able to grow on xylose as a sole carbon source. In contrast to aerobic glucose metabolism, S. cerevisiae XYL1-XYL2 transformants utilize xylose almost entirely oxidatively.
Kötter
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Institut für Mikrobiologie, Heinrich-Heine-Universität, Düsseldorf, Federal Republic of Germany.
Amore
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Hollenberg
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CP
Ciriacy
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D000429:Alcohol Oxidoreductases / Q000235:genetics / Q000378:metabolism; D000449:Aldehyde Reductase / Q000235:genetics; D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D050542:D-Xylulose Reductase; D015870:Gene Expression; D005800:Genes, Fungal; D008969:Molecular Sequence Data; D016366:Open Reading Frames; D010843:Pichia / Q000201:enzymology / Q000235:genetics*; D015183:Restriction Mapping; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000235:genetics*; D013401:Sugar Alcohol Dehydrogenases / Q000235:genetics*; D014170:Transformation, Genetic; D014994:Xylose / Q000378:metabolism
D014994:Xylose; D000429:Alcohol Oxidoreductases; D013401:Sugar Alcohol Dehydrogenases; C035795:D-xylose dehydrogenase; D000449:Aldehyde Reductase; D050542:D-Xylulose Reductase