De Santi Concetta, Ambrosino Luca, Tedesco Pietro, Zhai Lei, Zhou Cheng, Xue Yanfen, Ma Yanhe, de Pascale Donatella
Inst. of Protein Biochemistry, National Research Council, Naples, I-80131, Italy.
State Key Laboratory of Microbial Resources and National Engineering Laboratory for Industrial Enzymes, Inst. of Microbiology, CAS, Beijing, 100101, China.
Biotechnol Prog. 2015 Jul-Aug;31(4):890-9. doi: 10.1002/btpr.2096. Epub 2015 May 15.
A salt-tolerant esterase, designated H9Est, was identified from a metagenomic library of the Karuola glacier. H9Est gene comprised 1071 bp and encoded a polypeptide of 357 amino acids with a molecular mass of 40 kDa. Sequence analysis revealed that H9Est belonged to the family IV of bacterial lypolitic enzyme. H9Est was overexpressed in Escherichia coli and the purified enzyme showed hydrolytic activity towards p-nitrophenyl esters with carbon chain from 2 to 8. The optimal esterase activity was at 40°C and pH 8.0 and the enzyme retained its activity towards some miscible organic solvents such as polyethylene glycol. A three-dimensional model of H9Est revealed that S200, D294, and H324 formed the H9Est catalytic triad. Circular Dichroism spectra and molecular dynamic simulation indicated that the esterase had a wide denaturation temperature range and flexible loops that would be beneficial for H9Est performance at low temperatures while retaining heat-resistant features.
从卡若拉冰川宏基因组文库中鉴定出一种耐盐酯酶，命名为H9Est。H9Est基因由1071个碱基对组成，编码一个由357个氨基酸组成的多肽，分子量为40 kDa。序列分析表明，H9Est属于细菌脂肪分解酶家族IV。H9Est在大肠杆菌中过表达，纯化后的酶对碳链长度为2至8的对硝基苯酯具有水解活性。最佳酯酶活性温度为40°C，pH值为8.0，该酶对一些可混溶的有机溶剂如聚乙二醇仍保持活性。H9Est的三维模型显示，S200、D294和H324形成了H9Est催化三联体。圆二色光谱和分子动力学模拟表明，该酯酶具有较宽的变性温度范围和灵活的环结构，这有利于H9Est在低温下发挥作用，同时保留耐热特性。
