Research on Construction of a Whole-Cell Biocatalyst for Xylan Degradation
Research on Construction of a Whole-Cell Biocatalyst for Xylan Degradation
复制标题
木聚糖降解全细胞生物催化剂的构建研究
DOI：
10.4028/www.scientific.net/amr.347-353.2599
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发表时间：
2011
期刊：
影响因子：
--
通讯作者：
G. He
中科院分区：
文献类型：
--
作者：
J. Lu;M. Cui;S. Du;Lu Yang;Qin Guo;H. Ruan;G. He
内切-1，4-β-木聚糖酶（E.C.3.2.1.8）是糖苷水解酶家族中的一员。它能够将取代的木聚糖聚合物的主链水解成随机大小的片段。由于木聚糖酶的这种能力，它可以为木质纤维素的降解服务，并促进木聚糖的应用。酶的细胞表面展示是酵母中最具吸引力的应用之一。在构建木聚糖酶全细胞生物催化剂方面有很好的应用前景。为此，根据酿酒酵母（Saccharomyces cerevisiae）的密码子偏好性，优化合成了黑曲霉（Aspergillus尼日尔）BCC 14405内切1，4-β-木聚糖酶B（XylB）的cDNA序列。扩增编码半乳糖激酶（GAL 1）启动子、α交配因子1（MFα1）前原分泌信号、完全密码子优化的XylB、α凝集素C端320个氨基酸、乙醇脱氢酶（ADH 1）终止子和kanMX盒的基因，并将其克隆到YEplac 181中，构建以α凝集素为锚的细胞表面展示载体pGMAAK-XylB。将pGMAAK-XylB转化到S.啤酒。结果表明，XylB在S.啤酒。同时，利用上述元件构建了分泌表达质粒，并以α-凝集素作为对照菌株，对XylB的特性进行了研究。经2%半乳糖诱导48 h后，XylB的表达活性达到63 U/g干重。与重组分泌XylB相比，展示的XylB的最适pH从5变为6，最适温度从50 °C变为60 °C。
Endo-1,4-β-xylanase (E.C.3.2.1.8) is a family of glycoside hydrolase. It is capable of hydrolyzing the backbone of substituted xylan polymers into fragments of random size. Due to this ability, xylanase can serve the degradation of lignocellulose, and facilitate the application of xylan. Cell-surface display of enzymes is one of the most attractive applications in yeast. It is a promising utilization in constructing the whole-cell biocatalyst of xylanase. For this purpose, a cDNA sequence of endo-1,4-β-xylanase B (XylB) from Aspergillus niger BCC14405 was optimized and synthesized according to the codon bias of Saccharomyces cerevisiae. The genes encoding galactokinase (GAL1) promoter, α-mating factor 1 (MFα1) pre-pro secretion signal, fully codon-optimized XylB, the 320 amino acids of C terminal of α-agglutinin, alcohol dehydrogenase (ADH1) terminator and kanMX cassette were amplified and cloned into YEplac181 to construct a cell-surface display vector called pGMAAK-XylB with α-agglutinin as an anchor. Then pGMAAK-XylB was transformed into S. cerevisiae. The results show XylB was immobilized and actively expressed on S. cerevisiae. Meanwhile, a secretion expression plasmid was also constructed using the above elements except α-agglutinin as a control strain in the study of characteristic of XylB. After an induction of 48 h by 2% galactose, the activity of displayed XylB reached 63 U/g dry-cell weight. The optimal pH of displayed XylB has changed from 5 to 6 and the optimal temperature has changed from 50 °C to 60 °C, comparing to the recombinant secretion XylB.
影响因子：
4.4
作者：
Katahira, S;Fujita, Y;Kondo, A
通讯作者：
Kondo, A