School of Life Science and Technology, China Pharmaceutical University, Nanjing 210009, China.
Department of Pharmacy, Jiangs u Cancer Hospital, Nanjing 210009, China.
Int J Biol Macromol. 2018 Apr 1;109:329-337. doi: 10.1016/j.ijbiomac.2017.12.004. Epub 2017 Dec 9.
Novel thermostable amylase need to be continuously explored with the improvement of industrial requirements. A new acidophilic and thermostable amylase producing bacterium isolated from spring was identified as Bacillus strain on the basis of 16S rDNA. The amylase was purified by ammonium sulphate precipitation, gel chromatography and anion exchange chromatography. SDS-PAGE revealed that the enzyme was monomeric with a molecular weight of 58 kDa. The amylase exhibited optimal activity at pH 5.0 and temperature 100 °C. Then the enzyme showed high stability in pH ranges 4.0-10.0 and more than 90% of maximal activity was found from 20 °C to 80 °C. Apart from good stability toward SDS and non-ionic detergent, the purified enzyme exhibited high compatibility with some inhibitors such as urea and EDTA. The results demonstrated the stability of the enzyme in different organic solvents. Moreover, we determined the amylase gene, compared the structure with α-amylase BAA and BLA and found some thermostability determinants in our enzyme. Overall, presenting various properties were including high thermostability, Ca-independency, broad temperature and pH profiles, organic-solvent tolerance as well as excellent stability with detergents. Such characteristics have not been reported for this type of enzyme, and the α-amylase will be a suitable candidate in industrial fields.
随着工业需求的提高,需要不断探索新型热稳定淀粉酶。根据 16S rDNA,从温泉中分离到一种嗜酸耐热淀粉酶产生菌,鉴定为芽孢杆菌属。通过硫酸铵沉淀、凝胶层析和阴离子交换层析对淀粉酶进行纯化。SDS-PAGE 显示该酶为单体,分子量为 58 kDa。该酶在 pH 5.0 和温度 100°C 时表现出最佳活性。然后,该酶在 pH 4.0-10.0 范围内表现出很高的稳定性,在 20°C-80°C 之间发现超过 90%的最大活性。除了对 SDS 和非离子洗涤剂具有良好的稳定性外,纯化的酶还与一些抑制剂(如尿素和 EDTA)具有很高的兼容性。结果表明该酶在不同有机溶剂中的稳定性。此外,我们确定了淀粉酶基因,与α-淀粉酶 BAA 和 BLA 的结构进行比较,发现我们的酶中有一些耐热稳定性决定因素。总的来说,该酶具有高耐热性、Ca 独立性、宽温度和 pH 谱、有机溶剂耐受性以及与洗涤剂的出色稳定性等特性,这种酶尚未报道过,α-淀粉酶将是工业领域的合适候选者。
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