Discovery and characterization of novel D-xylose-specific transporters from Neurospora crassa and Pichia stipitis
Discovery and characterization of novel D-xylose-specific transporters from Neurospora crassa and Pichia stipitis
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DOI：
10.1039/c0mb00007h
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发表时间：
2010-01-01
影响因子：
--
通讯作者：
Zhao, Huimin
中科院分区：
文献类型：
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作者：
Du, Jing;Li, Sijin;Zhao, Huimin
酿酒酵母被认为是利用木质纤维素原料生产乙醇最有前途的生物之一。不幸的是，占木质纤维素高达30%的戊糖不能被野生型S利用。酿酒。将异源代谢途径引入S.酿酒酵母，使利用D-木糖，最丰富的戊糖。然而，所得到的重组S.酿酒酵母菌株在以D-木糖作为唯一碳源生长时表现出缓慢的生长速率和较差的糖利用效率。D-木糖的摄取是D-木糖利用的第一步。D-木糖只能通过己糖转运蛋白进入酵母细胞，与己糖相比，己糖转运蛋白对D-木糖的亲和力低两个数量级。研究还表明，在某些D-木糖代谢酵母菌株中，戊糖摄取效率低下是限制性步骤。在这里，我们报告了两个新的D-木糖特异性转运蛋白的克隆和表征粗糙脉孢菌和树干毕赤酵母。这两种转运蛋白是从总共18种假定的戊糖转运蛋白中鉴定出来的。它们在S.如分别通过HPLC分析和荧光共聚焦显微镜所指示的。使用C-14标记的糖摄取测定确定D-木糖特异性转运蛋白的动力学参数。戊糖特异性转运蛋白的使用应改善快速D-木糖同化菌株中的D-木糖消耗和乙醇生产，从而降低木质纤维素乙醇生产的成本。
Saccharomyces cerevisiae is considered one of the most promising organisms for ethanol production from lignocellulosic feedstock. Unfortunately, pentose sugars, which comprise up to 30% of lignocellulose, cannot be utilized by wild type S. cerevisiae. Heterologous pathways were introduced into S. cerevisiae to enable utilization of D-xylose, the most abundant pentose sugar. However, the resulting recombinant S. cerevisiae strains exhibited a slow growth rate and poor sugar utilization efficiency when grown on D-xylose as the sole carbon source. D-xylose uptake is the first step of D-xylose utilization. D-xylose can only enter yeast cells through hexose transporters, which have two orders of magnitude lower affinity towards D-xylose compared to hexoses. It was also shown that inefficient pentose uptake is the limiting step in some D-xylose metabolizing yeast strains. Here we report the cloning and characterization of two novel D-xylose-specific transporters from Neurospora crassa and Pichia stipitis. These two transporters were identified from a total of 18 putative pentose transporters. They were functionally expressed and properly localized in S. cerevisiae as indicated by HPLC analysis and fluorescence confocal microscopy, respectively. Kinetic parameters of the D-xylose-specific transporters were determined using a C-14-labeled sugar uptake assay. Use of pentose-specific transporters should improve D-xylose consumption and ethanol production in fast D-xylose assimilating strains, thereby lowering the cost of lignocellulosic ethanol production.