Tan Hao, Miao Renyun, Liu Tianhai, Cao Xuelian, Wu Xiang, Xie Liyuan, Huang Zhongqian, Peng Weihong, Gan Bingcheng
Soil and Fertilizer Institute, Sichuan Academy of Agricultural Sciences, Chengdu 610066, P.R. China.
Scientific Observing and Experimental Station of Agro-microbial Resource and Utilization in Southwest China, Ministry of Agriculture, Chengdu 610066, P.R. China.
J Microbiol Biotechnol. 2016 Oct 28;26(10):1717-1722. doi: 10.4014/jmb.1604.04051.
A novel phytase of Acidobacteria was identified from a soil metagenome, cloned, overexpressed, and purified. It has low sequence similarity (<44%) to all the known phytases. At the optimum pH (2.5), the phytase shows an activity level of 1,792 μmol/min/mg at physiological temperature (37°C) and could retain 92% residual activity after 30 min, indicating the phytase is acidophilic and acidostable. However the phytase shows poor stability at high temperatures. To improve its thermal resistance, the enzyme was redesigned using Disulfide by Design 2.0, introducing four additional disulfide bridges. The half-life time of the engineered phytase at 60°C and 80°C, respectively, is 3.0× and 2.8× longer than the wild-type, and its activity and acidostability are not significantly affected.
从土壤宏基因组中鉴定出一种新型嗜酸菌属植酸酶，进行了克隆、过表达和纯化。它与所有已知植酸酶的序列相似性较低（<44%）。在最佳pH值（2.5）下，该植酸酶在生理温度（37°C）时的活性水平为1792 μmol/分钟/毫克，30分钟后可保留92%的残余活性，表明该植酸酶嗜酸且耐酸。然而，该植酸酶在高温下稳定性较差。为提高其耐热性，使用设计二硫键2.0对该酶进行重新设计，引入了四个额外的二硫键。工程化植酸酶在60°C和80°C下的半衰期分别比野生型长3.0倍和2.8倍，其活性和耐酸性没有受到显著影响。
