Abstract:

Xyn11P13, a glycoside hydrolase family 11 xylanse gene, was cloned from Phialophora sp. P13. The full-length gene consists of 705 bp and encodes 234 amino acids, including a putative signal peptide of 19 residues at N-terminus. The mature xylanase was expressed in Pichia pastoris. In shaker flask culture, XYN11P13 was produced with activity of 11.8 U/mL. XYN11P13 showed the optimal activity at pH 5.5, it retained >80.0% of the maximal activity at pH 3.5~7.0, and showed high stability over the pH range 4.0~10.0. The optimum temperature was 65℃, it retained 80% of the maximal activity at 75℃, and was stable at 60℃. The specific activity, Km and Vmax values towards beechwood xylan were 137 U/mg, 6.9 mg/mL and 209 μmol/min/mg, respectively. XYN11P13 was strongly resistant to most metal ions. All these favorable properties make XYN11P13 attractive for potential applications in food and animal feed industries.

Key words: Phialophora sp.P13, Xylanase, cloning, Pichia pastoris, enzymatic properties

摘要：

从瓶霉Phialophora sp.P13菌株中克隆得到一个11家族的木聚糖酶基因,命名为xyn11P13。该基因ORF全长705 bp,共编码234个氨基酸和一个终止密码子,N端19个氨基酸为预测的信号肽序列。成熟蛋白在毕赤酵母中进行了重组表达,在摇床水平上木聚糖酶XYN11P13的表达量为11.8 U/mL。XYN11P13的最适pH为5.5,在pH 3.5~7.0可以保持80%以上的酶活性,在pH 4.0~10.0具有良好的pH稳定性。最适作用温度为65℃,并在60℃条件下保持稳定。以榉木木聚糖为底物,XYN11P13的比活,Km及Vmax值分别为205.3 U/mg,2.73 mg/mL,399.6 μmol/min/mg。多数金属离子对XYN11P13酶活力没有明显的影响。因此,XYN11P13具潜在优势应用于食品与动物饲料工业。

关键词: Phialophora sp. P13;木聚糖酶;克隆;毕赤酵母;酶学性质