A thermophilic and acid stable family-10 xylanase from the acidophilic fungus Bispora sp MEY-1
A thermophilic and acid stable family-10 xylanase from the acidophilic fungus Bispora sp MEY-1
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DOI：
10.1007/s00792-009-0272-0
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发表时间：
2009-09-01
期刊：
影响因子：
2.9
通讯作者：
Yao, Bin
中科院分区：
文献类型：
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作者：
Luo, Huiying;Li, Jiang;Yao, Bin
从嗜酸真菌Bisporasp. MEY-1中克隆了编码嗜热内切1，4 β-木聚糖酶（XYL 10 C）的完整基因xyl 10 C，并在毕赤酵母中表达。XYL 10 C与糖苷水解酶家族10的烟曲霉Af 293的推定木聚糖酶具有最高的核苷酸和氨基酸序列同一性，分别为57.3%和49.7%。在3.7-l发酵罐中实现了在巴斯德毕赤酵母中的高表达水平（73，400 U ml（-1））。纯化的重组XYL 10 C是嗜热的，在85 A ℃下表现出最大活性，这高于任何真菌木聚糖酶的报道。该酶也是高度热稳定的，表现出根类似于100%的初始活性后，在80 A摄氏度孵育60分钟，并在90 A摄氏度的活性> 87%，10分钟。XYL 10 C在80和85 A摄氏度的半衰期分别约为45和3小时。在pH3.0和4.5-5.0（最大值）有两个活性峰，且对酸非常稳定，在pH1.5 -6.0孵育1h后仍保持80%以上的活性。该酶对Co ~（2+）、Mn ~（2+）、Cr ~（3+）和Ag ~+具有较强的抗性。XYL 10 C对燕麦斯卑尔脱木聚糖的比活性为18，831 U mg（-1）。该酶具有广泛的底物特异性，以燕麦斯卑尔脱木聚糖为底物，转化产物简单（木糖65.1%，木二糖25.0%，木聚糖聚合物9.9%）。
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 85A degrees C, which is higher than that reported from any fungal xylanase. The enzyme was also highly thermostable, exhibiting root similar to 100% of the initial activity after incubation at 80A degrees C for 60 min and > 87% of activity at 90A degrees C for 10 min. The half lives of XYL10C at 80 and 85A 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 Co2+, Mn2+, Cr3+ 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.