Construction of whole-cell biocatalyst for xylan degradation through cell-surface xylanase display in Saccharomyces cerevisiae
Construction of whole-cell biocatalyst for xylan degradation through cell-surface xylanase display in Saccharomyces cerevisiae
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DOI：
10.1016/s1381-1177(02)00027-9
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发表时间：
2002-06-07
影响因子：
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通讯作者：
Kondo, A
中科院分区：
文献类型：
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作者：
Fujita, Y;Katahira, S;Kondo, A
我们构建了一种在细胞表面展示里氏木霉木聚糖酶H（XYNII）的酵母全细胞生物催化剂，并赋予酵母细胞降解木聚糖的能力。构建了编码XYNII成熟区和酵母α-凝集素C-末端一半（从C-末端算起320个氨基酸残基）的融合基因（XYNII-alpha-agglutinin），并在甘油醛-3-磷酸脱氢酶（GAPDH）启动子的控制下在酿酒酵母中表达。构建了RGSHis 6标记的XYNII-α-凝集素融合基因表达系统（RGSHis 6-XYNII-α-agglutinin）。免疫荧光标记确认RGSHis 6-XYNII-alpha-凝集素融合蛋白的细胞表面展示，并确认表达XYNU-alpha-凝集素和RGSHis 6-XYNII-alpha-凝集素但不在培养基中的酵母细胞中具有相似的木聚糖酶活性，表明XYNII以活性形式展示在细胞表面。展示XYNII的基于酵母的全细胞生物催化剂分别在pH 5.0和40 ℃下显示出最高的XYNII活性。这种全细胞生物催化剂不仅可以应用于木聚糖发酵生产乙醇的第一步，而且可以应用于低聚木糖的生产。(C)2002 Elsevier Science B. V.保留所有权利。
We constructed a yeast-based whole-cell biocatalyst displaying Trichoderma reesei xylanase H (XYNII) on the cell-surface and endowed the yeast-cells with the ability to degrade xylan. The fusion gene encoding the mature region of XYNII and the C-terminal half (320 amino acid residues from the C-terminal end) of yeast alpha-agglutinin (XYNII-alpha-agglutinin) was constructed and expressed in Saccharomyces cerevisiae under the control of a glyceraldehyde-3-phosphate dehydrogenase (GAPDH) promoter. The expression system of fusion gene encoding XYNII-alpha-agglutinin tagged with RGSHis6 consisting of arginine, glycine, serine, and histidine hexamer (RGSHis6-XYNII-alpha-agglutinin) was also constructed. Immunofluorescence labeling to confirm cell-surface display of the RGSHis6-XYNII-alpha-agglutinin fusion protein, and confirmation of similar xylanase activity in yeast-cells expressing XYNU-alpha-agglutinin and RGSHis6-XYNII-alpha-agglutinin but not in the culture medium, indicated that XYNII was displayed on the cell-surface in the active form. The XYNII-displaying yeast-based whole-cell biocatalyst showed highest XYNII activity at pH 5.0 and 40 degreesC, respectively. This whole-cell biocatalyst is expected to find application not only in the first step of fermentation of xylan to ethanol but also in xylooligosaccharide production. (C) 2002 Elsevier Science B.V. All rights reserved.