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首页> 外文期刊>Journal of Biotechnology >Functionality improvement of fungal lignin peroxidase by DNA shuffling for 2,4-dichlorophenol degradability and H(2)O(2) stability
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Functionality improvement of fungal lignin peroxidase by DNA shuffling for 2,4-dichlorophenol degradability and H(2)O(2) stability

机译：DNA改组为2,4-二氯苯酚可降解性和H（2）O（2）稳定性的真菌木质素过氧化物酶功能改进

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摘要

One of the major problems of wild-type lignin peroxidase (LiP) is its inactivity at the presence of excess H(2)O(2) and high concentration of aromatic compounds. Little is known about the substrate-binding site of LiP, and functionality improvement of LiP was not actively tried by genetic engineering and directed evolution. In order to improve LiPs functionality, we performed directed evolution with a colorimetric screening method. Finally, three types of LiP mutants were screened. The catalytic efficiency of the variants toward 2,4-dichlorophenol (DCP) degradation activity and the stability against H(2)O(2) was increased over the wild type. The K(m) value of the variants toward H(2)O(2) was increased, but K(m) value toward 2,4-DCP degradation was reduced. Overall, The K(cat)/K(m) values of the mutants toward 2,4-DCP was increased ca. 4-fold, and that toward H(2)O(2) was increased ca. 89-fold. Amino acid sequence analysis indicated that the most of the mutations were located on the enzyme surface. We expect that these results coupled with recombining mutation can be successfully applied to the molecular evolution cycles for screening of LiPs and other oxidative enzymes with improved functionality and stability.
机译：野生型木质素过氧化物酶（LiP）的主要问题之一是在过量H（2）O（2）和高浓度的芳香族化合物存在下其无活性。关于LiP的底物结合位点知之甚少，而且基因工程和定向进化并未积极地尝试改善LiP的功能。为了改善LiPs的功能，我们用比色筛选方法进行了定向进化。最后，筛选了三种类型的LiP突变体。变种对2，4-二氯苯酚（DCP）降解活性和对H（2）O（2）的稳定性的催化效率已超过野生型。朝向H（2）O（2）的变体的K（m）值增加，但是朝向2,4-DCP降解的K（m）值减小。总体而言，朝向2，4-DCP的突变体的K（cat）/ K（m）值增加了。 4倍，并向H（2）O（2）约增加。 89倍。氨基酸序列分析表明，大多数突变位于酶表面。我们希望将这些结果与重组突变相结合，可以成功地应用于分子进化周期中，以筛选具有改进的功能和稳定性的LiPs和其他氧化酶。

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