| Budget Amount *help |
¥14,200,000 (Direct Cost: ¥14,200,000)
Fiscal Year 1999: ¥5,600,000 (Direct Cost: ¥5,600,000)
Fiscal Year 1998: ¥8,600,000 (Direct Cost: ¥8,600,000)
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| Research Abstract |
Novel yeast cells armed with biocatalysts - glucoamylase, α-amylase, CM-cellulase, β-glucosidase, and lipase - were constructed by a cell surface engineering system of yeast Saccharomyces cerevisiae. These surface-engineered yeast cells were termed "Arming yeasts". The gene encoding Rhizopus oryzae glucamylase with its secretion signal peptide was fused with the gene encoding the C-terminal half of yeast α-agglutinin. Glucoamylase was shown to be displayed on the cell surface of S. cerevisiae in its active form, anchored covalently to the cell wall. S. cerevisiae is unable to utilize starch, while the arming cells could grow on starch as the sole carbon source. For enhancement of the ability to directly ferment starchy materials by the arming yeast, a surface-engineered yeast cell displaying two amylolytic enzymes was constructed. The gene encoding R. oryzae glucoamylase with its own secretion signal peptide and a truncated fragment of the a-amylase gene from Bacillus stearothermophilu
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s with the prepro secretion signal sequence of the yeast α-factor, respectively, were fused with the gene encoding the C-terminal half of the yeast α-agglutinin. The arming cell co-displaying glucoamylase and α-amylase could grow faster on starch as the sole carbon source than the cell displaying only glucoamylase. Furthermore, a novel cellulose-utilizing yeast cell displaying cellulolytic enzymes in their active forms on the cell surface of S. cerevisiae was constructed by the cell surface engineering. An arming yeast codisplaying FI-carboxymethylcellulase(CM-cellulase), one of the endo-type cellulase, and β-glucosidase from Aspergillus aculeatus was endowed with the ability of celloligosaccharide assimilation, suggesting the possibility that the assimilation of cellulosic materials may be carried out by S. cerevisiae expressing heterologous cellulase genes on the cell surface. Furthermore, a yeast cell armed with R. oryzae lipase was also constructed. The technique will be applied to endow cells with the ability of bioremediation. Less
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