Luo Huiying, Li Jiang, Yang Jun, Wang Hui, Yang Yuhui, Huang Huoqing, Shi Pengjun, Yuan Tiezheng, Fan Yunliu, Yao Bin
Key Laboratory for Feed Biotechnology of the Ministry of Agriculture, Feed Research Institute, Chinese Academy of Agricultural Sciences, No. 12 Zhongguancun South Street, 100081, Beijing, People's Republic of China.
Extremophiles. 2009 Sep;13(5):849-57. doi: 10.1007/s00792-009-0272-0. Epub 2009 Aug 5.
A complete gene, xyl10C, encoding a thermophilic endo-1,4-beta-xylanase (XYL10C), was cloned from the acidophilic fungus Bispora sp. MEY-1 and expressed in Pichia pastoris. XYL10C shares highest nucleotide and amino acid sequence identities of 57.3 and 49.7%, respectively, with a putative xylanase from Aspergillus fumigatus Af293 of glycoside hydrolase family 10. A high expression level in P. pastoris (73,400 U ml(-1)) was achieved in a 3.7-l fermenter. The purified recombinant XYL10C was thermophilic, exhibiting maximum activity at 85 degrees C, which is higher than that reported from any fungal xylanase. The enzyme was also highly thermostable, exhibiting approximately 100% of the initial activity after incubation at 80 degrees C for 60 min and >87% of activity at 90 degrees C for 10 min. The half lives of XYL10C at 80 and 85 degrees C were approximately 45 and 3 h, respectively. It had two activity peaks at pH 3.0 and 4.5-5.0 (maximum), respectively, and was very acid stable, retaining more than 80% activity after incubation at pH 1.5-6.0 for 1 h. The enzyme was resistant to Co(2+), Mn(2+), Cr(3+) and Ag(+). The specific activity of XYL10C for oat spelt xylan was 18,831 U mg(-1). It also had wide substrate specificity and produced simple products (65.1% xylose, 25.0% xylobiose and 9.9% xylan polymer) from oat spelt xylan.
从嗜酸真菌Bispora sp. MEY-1中克隆出一个完整的基因xyl10C，其编码嗜热内切-1,4-β-木聚糖酶（XYL10C），并在毕赤酵母中表达。XYL10C与烟曲霉Af293的推定木聚糖酶（糖苷水解酶家族10）的核苷酸和氨基酸序列一致性最高，分别为57.3%和49.7%。在3.7升发酵罐中，XYL10C在毕赤酵母中实现了高表达水平（73400 U ml(-1)）。纯化后的重组XYL10C具有嗜热性，在85℃时表现出最大活性，这高于任何报道的真菌木聚糖酶。该酶也具有高度热稳定性，在80℃孵育60分钟后仍保留约100%的初始活性，在90℃孵育10分钟后仍保留>87%的活性。XYL10C在80℃和85℃的半衰期分别约为45小时和3小时。它在pH 3.0和4.5 - 5.0（最大值）处分别有两个活性峰，并且非常耐酸，在pH 1.5 - 6.0孵育1小时后仍保留超过80%的活性。该酶对Co(2+)、Mn(2+)、Cr(3+)和Ag(+)具有抗性。XYL10C对燕麦 speltoid木聚糖的比活性为18831 U mg(-1)。它还具有广泛的底物特异性，从燕麦 speltoid木聚糖产生简单产物（65.1%木糖、25.0%木二糖和9.9%木聚糖聚合物）。
