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Isolation and Characterization of a New Thermoalkalophilic Lipase from Soil Bacteria

机译:土壤细菌中一种新型嗜热脂肪酶的分离与鉴定

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摘要

Lipases are diversified enzymes in their properties and substrate specificity, which make them attractive tools for various industrial applications. In this study, an alkalinethermostable lipase producing bacteria were isolated from soil of different regions of Isfahan province (Iran) and its lipase was purified by ammonium sulfate precipitation and ion exchange chromatography. To select a thermoalkalophil lipase producing bacterium, Rhodamine B and Horikoshi media were used and the strain that can grow at 45°Cwas selected. The isolated strain was identified using microbial and biochemical tests.One strain showed an orange colored zone on plate and grew on Horikoshi plate. Microbial and biochemical tests showed that the isolated strain was Bacillus subtilis, a Gram positive rod. In PCR, an expected band was obtained with about 371 bp. The activity of the purified lipase was 10.2 folds that of the standard enzyme using ammonium sulfate precipitation and ion exchange chromatography. The molecular weight of lipase determined by SDS-PAGE electrophoresis, was 21 and 35 KDa.Existence of two bands in SDS-PAGE electrophoresis and low amount of obtained purified enzyme highlights the necessity of optimization of purification and concentrating process.
机译:脂肪酶在性质和底物特异性方面是多种酶,这使其成为用于各种工业应用的有吸引力的工具。在这项研究中,从伊斯法罕省(伊朗)不同地区的土壤中分离出了碱性热稳定脂肪酶产生细菌,并通过硫酸铵沉淀和离子交换色谱法纯化了其脂肪酶。为了选择产热嗜碱脂肪酶的细菌,使用了若丹明B和Horikoshi培养基,并选择了可以在45℃生长的菌株。分离的菌株通过微生物和生化测试鉴定。一种菌株在平板上显示橙色区域,并在Horikoshi平板上生长。微生物和生化测试表明,分离出的菌株是枯草芽孢杆菌,一种革兰氏阳性棒。在PCR中,获得了预期的约371bp的条带。使用硫酸铵沉淀和离子交换色谱法,纯化的脂肪酶的活性是标准酶的10.2倍。通过SDS-PAGE电泳测定的脂肪酶分子量分别为21和35KDa。SDS-PAGE电泳中存在两个条带,且所获得的纯化酶量很少,这突出表明有必要对纯化和浓缩工艺进行优化。

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