Graduate School of Bioresources, Mie University, Tsu, 514-8507, Japan.
Enzyme Microb Technol. 2019 May;124:23-31. doi: 10.1016/j.enzmictec.2019.01.011. Epub 2019 Jan 30.
The abnA gene from Ruminiclostridium josui encodes the large modular arabinanolytic enzyme, Abf43A-Abf43B-Abf43C, consisting of an N-terminal signal peptide, a Laminin_G_3 module, a GH43_22 module, a Laminin_G_3 module, a Big_4 module, a GH43_26 module, a GH43_34 module and a dockerin module in order with a calculated molecular weight of 204,108. Three truncated enzymes were recombinantly produced in Escherichia coli and biochemically characterized, RjAbf43A consisting of the first Laminin_G_3 module and GH43_22 module, RjAbf43B consisting of the second Laminin_G_3 module, Big_4 module and GH43_26 module, and RjAbf43C consisting of the GH43_34 module. RjAbf43A showed a strong α-l-arabinofuranosidase activity toward sugar beet arabinan, highly branched arabinan but not linear arabinan, thus it acted in the removal of arabinose side chains from sugar beet arabinan. By contrast, RjAbf43B showed a strong exo-α-1,5-l-arabinofuranosidase activity toward linear arabinan and arabinooligosaccharides whereas RjAbf43C showed low activity toward these substrates. Although RjAbf43B was activated by the presence of some metal ions such as Zn, Mg and Ni, RjAbf43A was inhibited by these ions. RjAbf43A and RjAbf43B attacked sugar beet arabinan in a synergistic manner. By comparison, RjAbf43A-Abf43B containing both GH43_22 and GH43_26 modules showed lower hydrolytic activity toward sugar beet arabinan but higher activity toward sugar beet fiber than the sum of the individual activities of RjAbf43A and RjAbf43B, suggesting that the coexistence of two distinct GH43 modules in a single polypeptide is important for the efficient hydrolysis of an insoluble and natural polysaccharide but not a soluble substrate.
来自鲁米努克利氏梭菌的 abnA 基因编码大型模块化阿拉伯聚糖酶 Abf43A-Abf43B-Abf43C，由一个 N 端信号肽、一个 Laminin_G_3 模块、一个 GH43_22 模块、一个 Laminin_G_3 模块、一个 Big_4 模块、一个 GH43_26 模块、一个 GH43_34 模块和一个 dockerin 模块组成，计算分子量为 204108。三种截短的酶在大肠杆菌中重组表达并进行了生化特性分析，RjAbf43A 由第一个 Laminin_G_3 模块和 GH43_22 模块组成，RjAbf43B 由第二个 Laminin_G_3 模块、Big_4 模块和 GH43_26 模块组成，RjAbf43C 由 GH43_34 模块组成。RjAbf43A 对糖甜菜阿拉伯聚糖、高度支化的阿拉伯聚糖具有很强的α-l-阿拉伯呋喃糖苷酶活性，但对线性阿拉伯聚糖没有活性，因此它可以从糖甜菜阿拉伯聚糖中去除阿拉伯糖侧链。相比之下，RjAbf43B 对线性阿拉伯聚糖和阿拉伯寡糖具有很强的外切-α-1,5-l-阿拉伯呋喃糖苷酶活性，而 RjAbf43C 对这些底物的活性较低。尽管 RjAbf43B 的活性受到 Zn、Mg 和 Ni 等一些金属离子的激活，但 RjAbf43A 则受到这些离子的抑制。RjAbf43A 和 RjAbf43B 以协同方式攻击糖甜菜阿拉伯聚糖。相比之下，含有 GH43_22 和 GH43_26 两个模块的 RjAbf43A-Abf43B 对糖甜菜阿拉伯聚糖的水解活性较低，但对糖甜菜纤维的活性高于 RjAbf43A 和 RjAbf43B 各自活性的总和，这表明单一多肽中两个不同 GH43 模块的共存对于高效水解不溶性天然多糖但不是可溶性底物很重要。
