Zverlov V, Piotukh K, Dakhova O, Velikodvorskaya G, Borriss R
Institut für Biologie, Humboldt Universität, Berlin, Germany.
Appl Microbiol Biotechnol. 1996 Mar;45(1-2):245-7. doi: 10.1007/s002530050678.
The nucleotide sequence of the xynA gene, encoding extracellular xylanase A of Thermotoga neapolitana, was determined. The xynA gene was 3264 base pairs (bp) long and encoded a putative polypeptide of 1055 amino acids. Three different domains were identified by sequence comparison and functional analysis of proteins with N- and/or C-terminal deletions. The core domain displayed significant homology to members of the glycosyl hydrolase family 10. N- and C-terminal domains were dispensable for enzymatic activity and seemed to be responsible for thermostability and cellulose binding, respectively. The intact gene and its truncated variants were expressed in Escherichia coli and purified for biochemical characterization. The enzyme was shown to act as an endo-1,4-beta-xylanase, but minor activities against lichenan, barley glucan, methylumbelliferyl cellobioside and p-nitrophenyl xyloside were also detected. The specific activity and pH and temperature optima for hydrolysis of oat xylan were 111.3 U.mg-1, 5.5 and 102 degrees C, respectively. The endoxylanase was stable at 90 degrees C and retained 50% activity when incubated for 2 h at 100 degrees C.
测定了编码嗜热栖热菌胞外木聚糖酶A的xynA基因的核苷酸序列。xynA基因长3264个碱基对（bp），编码一个推定的含1055个氨基酸的多肽。通过对具有N端和/或C端缺失的蛋白质进行序列比较和功能分析，确定了三个不同的结构域。核心结构域与糖基水解酶家族10的成员具有显著同源性。N端和C端结构域对酶活性不是必需的，似乎分别负责热稳定性和纤维素结合。完整基因及其截短变体在大肠杆菌中表达并纯化以进行生化特性分析。该酶被证明可作为内切-1,4-β-木聚糖酶，但也检测到其对地衣多糖、大麦葡聚糖、甲基伞形酮纤维二糖苷和对硝基苯基木糖苷有轻微活性。燕麦木聚糖水解的比活性、最适pH和温度分别为111.3 U.mg-1、5.5和102℃。该内切木聚糖酶在90℃下稳定，在100℃下孵育2小时后仍保留50%的活性。
