Zhang Zhiwei, Dong Junshuai, Zhang Deqing, Wang Jiaojiao, Qin Xing, Liu Bo, Xu Xinxin, Zhang Wei, Zhang Yuhong
College of Forestry, Shanxi Agricultural University, Taigu, Shanxi 030801, China.
Biotechnology Research Institute, Chinese Academy of Agricultural Sciences, No. 12 Zhongguancun South Street, Beijing 100081, China.
J Biosci Bioeng. 2018 Dec;126(6):690-696. doi: 10.1016/j.jbiosc.2018.05.022. Epub 2018 Jul 3.
A pectin methylesterase gene, pme-zj5a, from Aspergillus niger ZJ5 was cloned and high-level expressed in Pichia pastoris. The highest PME activity was 71.11 U/ml after induction with methanol for 20 h at 30 °C. The molecular mass of purified PME-ZJ5A was estimated to be 37 kDa by SDS-PAGE, and its K, V and k values of PME-ZJ5A were determined to be 3.27 mg/ml, 5.36 μmol/min/mg, and 22.33 s with pectin. Purified recombinant PME-ZJ5A exhibited optimal activity at pH 3.8 and 45 °C. It retained more than 60% of its maximum activity at 10 °C. Moreover, recombinant PME-ZJ5A can increase the transmittance of pineapple juice by 60.8%, and increase the firmness of pineapple cubes nearly double when combined with CaCl, which showed good potential in fruit processing.
从黑曲霉ZJ5中克隆了一个果胶甲酯酶基因pme-zj5a，并在毕赤酵母中进行了高水平表达。在30℃用甲醇诱导20小时后，最高PME活性为71.11 U/ml。通过SDS-PAGE估计纯化的PME-ZJ5A的分子量为37 kDa，其对果胶的Km、Vmax和kcat值分别为3.27 mg/ml、5.36 μmol/min/mg和22.33 s。纯化的重组PME-ZJ5A在pH 3.8和45℃时表现出最佳活性。在10℃时，它保留了超过60%的最大活性。此外，重组PME-ZJ5A可使菠萝汁的透光率提高60.8%，与氯化钙结合时可使菠萝块的硬度增加近一倍，在水果加工中显示出良好的潜力。
