构建具有油水界面独立性的黑曲霉脂肪酶突变体。
Construction of Aspergillus niger lipase mutants with oil-water interface independence.
机构信息
Engineering Research Center of Industrial Microbiology, Ministry of Education, Fujian Normal University, Fuzhou, Fujian 350108, PR China.
出版信息
Enzyme Microb Technol. 2011 Feb 8;48(2):129-33. doi: 10.1016/j.enzmictec.2010.10.011. Epub 2010 Nov 9.
Based on previous bioinformational analytical results [Shu ZY, et al. Biotechnol Prog 2009;25:409-16], four A. niger lipase (ANL) mutants, ANL-Ser84Gly, ANL-Asp99Pro, ANL-Lys108Glu and ANL-EαH (obtained by replacing the lid domain of ANL with the corresponding domain from A. niger feruloyl esterase), were constructed to screen out ANL mutants with oil-water interface independence. ANL-S84G displayed a pronounced interfacial activation, while ANL-D99P and ANL-K108E displayed no interfacial activation. The specific activity of ANL-S84G towards p-nitrophenyl esters decreased from 29.8% to 76.5% compared with that of ANL, while the specific activity of ANL-D99P towards p-nitrophenyl palmitate increased 2.2-fold. The thermostability of ANL-K108E was almost unchanged, while the thermostability of ANL-S84G and ANL-D99P significantly decreased compared with that of ANL. The construction of oil-water interface-independent ANL mutants would help to further understand the mechanism of lipase interfacial activation.
基于先前的生物信息学分析结果[Shu ZY, et al. Biotechnol Prog 2009;25:409-16]，构建了四个黑曲霉脂肪酶（ANL）突变体，即 ANL-Ser84Gly、ANL-Asp99Pro、ANL-Lys108Glu 和 ANL-EαH（通过用来自黑曲霉阿魏酸酯酶的相应结构域替换 ANL 的盖结构域获得），以筛选出具有油水界面独立性的 ANL 突变体。ANL-S84G 表现出明显的界面激活，而 ANL-D99P 和 ANL-K108E 则没有表现出界面激活。与 ANL 相比，ANL-S84G 对 p-硝基苯酯的比活性从 29.8%降低到 76.5%，而 ANL-D99P 对 p-硝基苯棕榈酸酯的比活性增加了 2.2 倍。ANL-K108E 的热稳定性几乎不变，而 ANL-S84G 和 ANL-D99P 的热稳定性与 ANL 相比则显著降低。构建油水界面非依赖性 ANL 突变体有助于进一步了解脂肪酶界面激活的机制。
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