Abstract：
Thermophilic Geobacillus kaustophilus HTA426 genome possesses a monoacylglycerol lipase (MAGL) gene. MAGLs can synthesize emulsifiers for use in the food and pharmaceutical industries from fatty acids and glycerol. They can also be used to analyze monoacylglycerol (MAG) levels in serum and food. The MAGL gene from strain HTA426 was artificially synthesized and heterologously expressed in Escherichia coli BL21(DE3). The recombinant His-tag fused MAGL (GkMAGL) was purified using a Ni2+-affinity column. The purified enzyme showed a temperature optimum at 65 °C and was stable up to 75 °C after 30 min incubation. In addition, the enzyme exhibited a pH optimum of 7.5 and was stable from pH 5.0 to 11.0. The enzyme hydrolyzed monoacylglycerols and showed the highest activity toward 1-monolauroylglycerol. The enzyme was stable in the presence of various organic solvents and detergents. The addition of Triton X-100 significantly increased GkMAGL activity. The thermal stability of the enzyme was higher than that of thermostable MAGL from Geobacillus sp. 12AMOR1 (12AMOR1_MAGL). Circular dichroism spectral analysis showed that the conformational stability of the GkMAGL was higher than that of 12AMOR1_MAGL at higher temperatures. These results indicate that the GkMAGL has useful features that can be used for various biotechnological applications.
摘要：
嗜热考斯特氏菌HTA426基因组具有单酰基甘油脂肪酶（MAGL）基因。MAGL可以从脂肪酸和甘油合成用于食品和制药工业的乳化剂。它们还可用于分析血清和食物中的单酰基甘油(MAG)水平。来自菌株HTA426的MAGL基因是人工合成的，并在大肠杆菌BL21（DE3）中异源表达。使用Ni2+亲和柱纯化重组His-标签融合的MAGL(GkMAGL)。纯化的酶在65°C显示出最佳温度，并且在孵育30分钟后在75°C下稳定。此外,该酶的最适pH为7.5，在pH5.0至11.0之间稳定。该酶水解单酰基甘油，对1-单月桂酰甘油的活性最高。该酶在各种有机溶剂和洗涤剂的存在下是稳定的。TritonX-100的添加显著增加了GkMAGL活性。该酶的热稳定性高于来自地芽孢杆菌的热稳定MAGL。12AMOR1(12AMOR1_MAGL)。圆二色光谱分析表明，在较高温度下，GkMAGL的构象稳定性高于12AMOR1_MAGL。这些结果表明GkMAGL具有可用于各种生物技术应用的有用特征。
