Tan Hao, Wu Xiang, Xie Liyuan, Huang Zhongqian, Peng Weihong, Gan Bingcheng
Soil and Fertilizer Institute, Sichuan Academy of Agricultural Sciences, Chengdu, China.
Scientific Observing and Experiment Station of Southwestern Region for Agricultural Microbial Resource Utilization, Ministry of Agriculture, Chengdu, China.
Appl Microbiol Biotechnol. 2016 Mar;100(5):2225-41. doi: 10.1007/s00253-015-7097-9. Epub 2015 Nov 4.
Phytases are enzymes degrading phytic acid and thereby releasing inorganic phosphate. While the phytases reported to date are majorly from culturable microorganisms, the fast-growing quantity of publicly available metagenomic data generated in the last decade has enabled bioinformatic mining of phytases in numerous data mines derived from a variety of ecosystems throughout the world. In this study, we are interested in the histidine acid phosphatase (HAP) family phytases present in insect-cultivated fungus gardens. Using bioinformatic approaches, 11 putative HAP phytase genes were initially screened from 18 publicly available metagenomes of fungus gardens and were further overexpressed in Escherichia coli. One phytase from a south pine beetle fungus garden showed the highest activity and was then chosen for further study. Biochemical characterization showed that the phytase is mesophilic but possesses strong ability to withstand high temperatures. To our knowledge, it has the longest half-life time at 100 °C (27 min) and at 80 °C (2.1 h) as compared to all the thermostable phytases publicly reported to date. After 100 °C incubation for 15 min, more than 93 % of the activity was retained. The activity was 3102 μmol P/min/mg at 37 °C and 4135 μmol P/min/mg at 52.5 °C, which is higher than all the known thermostable phytases. For the high activity level demonstrated at mesophilic temperatures as well as the high resilience to high temperatures, the phytase might be promising for potential application as an additive enzyme in animal feed.
植酸酶是降解植酸从而释放无机磷酸盐的酶。虽然迄今为止报道的植酸酶主要来自可培养微生物，但过去十年中快速增长的公开宏基因组数据量使得能够在来自世界各地各种生态系统的众多数据宝库中对植酸酶进行生物信息学挖掘。在本研究中，我们对昆虫培养的真菌园中的组氨酸酸性磷酸酶（HAP）家族植酸酶感兴趣。使用生物信息学方法，最初从18个公开可用的真菌园宏基因组中筛选出11个推定的HAP植酸酶基因，并在大肠杆菌中进一步过表达。来自南方松甲虫真菌园的一种植酸酶表现出最高活性，随后被选作进一步研究。生化特性表明，该植酸酶是嗜温的，但具有很强的耐高温能力。据我们所知，与迄今为止公开报道的所有耐热植酸酶相比，它在100℃（27分钟）和80℃（2.1小时）下具有最长的半衰期。在100℃孵育15分钟后，仍保留超过93%的活性。该酶在37℃时的活性为3102μmol P/分钟/毫克，在52.5℃时为4135μmol P/分钟/毫克，高于所有已知的耐热植酸酶。由于该植酸酶在嗜温温度下表现出高活性水平以及对高温具有高耐受性，它可能有望作为动物饲料中的添加酶进行潜在应用。
