Tunnel engineering for modulating the substrate preference in cytochrome P450BsβHI
Tunnel engineering for modulating the substrate preference in cytochrome P450BsβHI
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DOI：
10.1186/s40643-021-00379-1
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发表时间：
2021-04-03
影响因子：
4.6
通讯作者：
Liu, Luo
中科院分区：
文献类型：
--
作者：
Meng, Shuaiqi;An, Ruipeng;Liu, Luo
活性位点通常位于酶的内部，因此底物应该通过隧道到达活性位点。隧道工程是改善酶催化性能的有力策略。在此，选择来自枯草芽孢杆菌的羟化酶P450（Bs beta）的P450（Bs beta）HI（Q85 H/V170 I）作为研究模型，其被报道为潜在的脱羧酶。然而，这种酶对长链脂肪酸表现出低脱羧酶活性。在此，进行隧道工程活动以调节底物偏好并改善P450（Bs beta）HI的脱羧活性。最终获得的Bs β HI-F79 A变体对棕榈酸的转化率提高了15.2倍; Bs β HI-F173 V变体对十五烷酸的转化率提高了3.9倍。这项研究表明，衬底的偏好可以调制隧道工程策略。
An active site is normally located inside enzymes, hence substrates should go through a tunnel to access the active site. Tunnel engineering is a powerful strategy for refining the catalytic properties of enzymes. Here, P450(Bs beta)HI (Q85H/V170I) derived from hydroxylase P450(Bs beta) from Bacillus subtilis was chosen as the study model, which is reported as a potential decarboxylase. However, this enzyme showed low decarboxylase activity towards long-chain fatty acids. Here, a tunnel engineering campaign was performed for modulating the substrate preference and improving the decarboxylation activity of P450(Bs beta)HI. The finally obtained Bs beta HI-F79A variant had a 15.2-fold improved conversion for palmitic acid; Bs beta HI-F173V variant had a 3.9-fold improved conversion for pentadecanoic acid. The study demonstrates how the substrate preference can be modulated by tunnel engineering strategy.