Coexpression of the Bacillus pumilus beta-xylosidase (xynB) gene with the Trichoderma reesei beta xylanase 2 (xyn2) gene in the yeast Saccharomyces cerevisiae.
The xynB gene encoding the Bacillus pumilus beta-xylosidase was expressed separately and jointly with the Trichoderma reesei beta-xylanase (xyn2) gene in the yeast Saccharomyces cerevisiae. Both genes were placed under the transcriptional control of the glucose-derepressible alcohol dehydrogenase 2 promoter (ADH2p) and terminator (ADH2T) sequences. The xynB gene was fused in frame to the yeast mating factor alpha1 secretion sequence (MFalpha1s) to effect secretion in S. cerevisiae. The fusion protein was designated Xlo1. Xlo1 produced in S. cerevisiae exhibited low affinity for xylobiose, but eventually hydrolyzed xylobiose and xylotriose to the monomeric constituent, D-xylose. Coproduction of Xyn2 and Xlo1 by S. cerevisiae led to a 25% increase in the amount of reducing sugars released from birchwood xylan compared to S. cerevisiae producing only the Xyn2 beta-xylanase. However, no D-xylose was produced from birchwood xylan, presumably due to very low Xlo1 beta-xylosidase activity and its low affinity for xylobiose.
La Grange
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DC
Department of Microbiology, University of Stellenbosch, South Africa.
Claeyssens
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Pretorius
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Van Zyl
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WH
D000426:Alcohol Dehydrogenase / Q000235:genetics; D001407:Bacillus / Q000201:enzymology / Q000235:genetics*; D015152:Blotting, Northern; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D043364:Endo-1,4-beta Xylanases; D015870:Gene Expression; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000235:genetics*; D017422:Sequence Analysis, DNA; D013728:Terminator Regions, Genetic; D014242:Trichoderma / Q000201:enzymology / Q000235:genetics*; D014990:Xylans / Q000378:metabolism; D014994:Xylose / Q000378:metabolism; D014995:Xylosidases / Q000235:genetics* / Q000378:metabolism
D012333:RNA, Messenger; D011994:Recombinant Proteins; D014990:Xylans; D014994:Xylose; D000426:Alcohol Dehydrogenase; D014995:Xylosidases; C026579:exo-1,4-beta-D-xylosidase; D043364:Endo-1,4-beta Xylanases