An Engineered Alcohol Oxidase for the Oxidation of Primary Alcohols
An Engineered Alcohol Oxidase for the Oxidation of Primary Alcohols
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DOI：
10.1002/cbic.201800556
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发表时间：
2019-01-18
期刊：
影响因子：
3.2
通讯作者：
Turner, Nicholas J.
中科院分区：
文献类型：
--
作者：
Heath, Rachel S.;Birmingham, William R.;Turner, Nicholas J.
以己-1-醇氧化为己醛为目标反应，对胆碱氧化酶进行了结构导向的定向进化。与野生型酶相比，鉴定出具有20倍增加的k（cat）的六个氨基酸的变体。该变体能够在不到24小时内将10 mm己醇氧化成己醛，转化率为100%。此外，该变体显示出热稳定性的显著增加，T-m相应增加20 ° C。用包括乙酸乙酯、庚烷和环己烷的有机溶剂证明了改进的溶剂耐受性，从而能够使醛的转化率比无溶剂体系的转化率高至多30%。尽管胆碱氧化酶向己-1-醇进化，但这种新变体也显示出对约50种主要脂肪族、不饱和、支链、环状、苄基和卤代醇的比活性增加（高达100倍）。
Structure-guided directed evolution of choline oxidase has been carried out by using the oxidation of hexan-1-ol to hexanal as the target reaction. A six-amino-acid variant was identified with a 20-fold increased k(cat) compared to that of the wild-type enzyme. This variant enabled the oxidation of 10 mm hexanol to hexanal in less than 24 h with 100 % conversion. Furthermore, this variant showed a marked increase in thermostability with a corresponding increase in T-m of 20 degrees C. Improved solvent tolerance was demonstrated with organic solvents including ethyl acetate, heptane and cyclohexane, thereby enabling improved conversions to the aldehyde by up to 30 % above conversion for the solvent-free system. Despite the evolution of choline oxidase towards hexan-1-ol, this new variant also showed increased specific activities (by up to 100-fold) for around 50 primary aliphatic, unsaturated, branched, cyclic, benzylic and halogenated alcohols.