耐盐嗜热菌 Thermobifida halotolerans YIM 90462 T 碱性耐热 GH9 内切葡聚糖酶的克隆、表达和特性分析。
Cloning, expression and characterization of an alkaline thermostable GH9 endoglucanase from Thermobifida halotolerans YIM 90462 T.
机构信息
Key Laboratory of Microbial Diversity in Southwest China, Ministry of Education and Laboratory for Conservation and Utilization of Bio-Resources, Yunnan Institute of Microbiology, Yunnan University, Kunming 650091, PR China.
出版信息
Bioresour Technol. 2011 Nov;102(21):10143-6. doi: 10.1016/j.biortech.2011.08.019. Epub 2011 Aug 10.
The endoglucanase gene, thcel9A, from Thermobifida halotolerans YIM 90462(T) was cloned and expressed in Escherichia coli BL 21(DE). The 2895-bp full-length gene encodes a 964-residue polypeptide (Thcel9A) containing a catalytic domain belonging to glycosyl hydrolases (GH) family 9. Phylogenetic analysis indicated that Thcel9A is closely related to Cel9A of Thermobifidafusca YX. Thcel9A was purified from the culture supernatant by Ni(2+)-affinity chromatography and the purified enzyme exhibited optimal activity at 55°C and pH 8.0. Substrate specificity assays showed that it not only had CMCase activity, but also hydrolase activity on microcrystalline cellulose and filter paper. These properties suggested that Thcel9A is a classical GH9 group A endoglucanase.
来自耐盐嗜热菌（Thermobifida halotolerans）YIM 90462(T)的内切葡聚糖酶基因 thcel9A 被克隆并在大肠杆菌 BL 21(DE)中表达。2895bp 的全长基因编码一个 964 个残基的多肽（Thcel9A），其中包含一个属于糖苷水解酶（GH）家族 9 的催化结构域。系统发育分析表明，Thcel9A 与嗜热纤维单胞菌（Thermobifidafusca YX）的 Cel9A 密切相关。Thcel9A 从培养上清液中通过 Ni(2+)-亲和层析进行纯化，纯化后的酶在 55°C 和 pH 8.0 下表现出最佳活性。底物特异性测定表明，它不仅具有 CMCase 活性，而且对微晶纤维素和滤纸具有水解酶活性。这些特性表明 Thcel9A 是一种典型的 GH9 组 A 内切葡聚糖酶。
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