Molecular investigation of a novel thermostable glucan phosphorylase from Thermoanaerobacter tengcongensis
Molecular investigation of a novel thermostable glucan phosphorylase from Thermoanaerobacter tengcongensis
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DOI：
10.1016/j.enzmictec.2007.03.008
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发表时间：
2007-08-02
影响因子：
3.4
通讯作者：
Xiang, Hua
中科院分区：
文献类型：
--
作者：
Chen, Shiqiong;Liu, Jingfang;Xiang, Hua
Thermoanaerobacter tengcongensis 中的 glgP 基因编码一种假定的 a-葡聚糖磷酸化酶 (Tte-GlgP)，该酶比大多数其他细菌中的同源物小得多，并且属于一个不太被认可的家族。为了表征这种新型酶，它在大肠杆菌中过表达并纯化至明显的同质性。这种纯化的 Tte-GlgP 具有预期的分子大小（类似于 64 kDa）和等电点（类似于 pI 6.2），在将多种葡聚糖（例如可溶性淀粉、麦芽糖糊精和糖原）转化为葡萄糖-1-磷酸时表现出广泛的底物谱。 Western blot分析表明，Tte-GIgP的产生受到葡萄糖的抑制，但受到麦芽糖的轻微诱导，表明它参与了该嗜热菌的碳水化合物代谢。 Tte-GlgP 在 60°C 时具有最佳活性，并且在 60°C 处理 6 h 后具有热稳定性，残留活性为 90%，因此过表达的 Tte-GlgP 也可能是其家族结构功能分析的良好候选者。 (c) 2007 Elsevier Inc. 保留所有权利。
The glgP gene in Thermoanaerobacter tengcongensis encodes a putative a-glucan phosphorylase (Tte-GlgP), which is much smaller than its homologs in most other bacteria and is grouped into a less-recognized family. To characterize this novel enzyme, it was overexpressed in Escherichia coli and purified to apparent homogeneity. This purified Tte-GlgP, with expected molecular size (similar to 64 kDa) and isoelectric point (similar to pI 6.2), exhibited a wide substrate spectrum in transformation of a variety of glucans such as soluble starch, maltodextrins and glycogen into glucose-1-phosphate. Western blot analysis indicated that the production of Tte-GIgP in T tengcongensis was repressed by glucose but induced by maltose slightly, suggesting that it was involved in the carbohydrate metabolism in this thermophile. The Tte-GlgP was optimally active at 60 degrees C, and was thermostable with 90% residual activity after treating at 60 degrees C for 6 h, thus the overexpressed Tte-GIgP might also be a good candidate for structure-function analysis in its family. (c) 2007 Elsevier Inc. All rights reserved.