Faculty of Science, Universiti Teknologi Malaysia, Skudai 81310, Johor, Malaysia.
Centre for Novel Agricultural Products, Department of Biology, University of York, Wentworth Way, York YO10 5DD, UK.
Int J Mol Sci. 2019 May 9;20(9):2284. doi: 10.3390/ijms20092284.
A halo-thermophilic bacterium, strain RA (previously known as bacterium RA), was isolated from a hot spring in Langkawi, Malaysia. A complete genome analysis showed that the bacterium harbors 57 glycoside hydrolases (GHs), including a multi-domain xylanase (XynRA2). The full-length XynRA2 of 813 amino acids comprises a family 4_9 carbohydrate-binding module (CBM4_9), a family 10 glycoside hydrolase catalytic domain (GH10), and a C-terminal domain (CTD) for type IX secretion system (T9SS). This study aims to describe the biochemical properties of XynRA2 and the effects of CBM truncation on this xylanase. XynRA2 and its CBM-truncated variant (XynRA2ΔCBM) was expressed, purified, and characterized. The purified XynRA2 and XynRA2ΔCBM had an identical optimum temperature at 70 °C, but different optimum pHs of 8.5 and 6.0 respectively. Furthermore, XynRA2 retained 94% and 71% of activity at 4.0 M and 5.0 M NaCl respectively, whereas XynRA2ΔCBM showed a lower activity (79% and 54%). XynRA2 exhibited a turnover rate () of 24.8 s, but this was reduced by 40% for XynRA2ΔCBM. Both the xylanases hydrolyzed beechwood xylan predominantly into xylobiose, and oat-spelt xylan into a mixture of xylo-oligosaccharides (XOs). Collectively, this work suggested CBM4_9 of XynRA2 has a role in enzyme performance.
一株嗜热 halo 菌，菌株 RA（先前称为细菌 RA），从马来西亚兰卡威的温泉中分离得到。全基因组分析表明，该细菌含有 57 种糖苷水解酶（GHs），包括一种多结构域木聚糖酶（XynRA2）。全长 813 个氨基酸的 XynRA2 包含一个家族 4_9 碳水化合物结合模块（CBM4_9）、一个家族 10 糖苷水解酶催化结构域（GH10）和一个用于九型分泌系统（T9SS）的 C 端结构域（CTD）。本研究旨在描述 XynRA2 的生化特性以及 CBM 截断对这种木聚糖酶的影响。表达、纯化和表征了 XynRA2 和其 CBM 截断变体（XynRA2ΔCBM）。纯化的 XynRA2 和 XynRA2ΔCBM 的最适温度相同，为 70°C，但最适 pH 分别为 8.5 和 6.0。此外，XynRA2 在 4.0 M 和 5.0 M NaCl 下分别保留了 94%和 71%的活性，而 XynRA2ΔCBM 的活性较低（79%和 54%）。XynRA2 的周转率（）为 24.8 s，但 XynRA2ΔCBM 的周转率降低了 40%。两种木聚糖酶均主要将桦木木聚糖水解成木二糖，并将燕麦-斯佩尔特木聚糖水解成混合的木低聚糖（XOs）。总的来说，这项工作表明 XynRA2 的 CBM4_9 在酶性能中起作用。
