Engineering of a Plant Isoprenyl Diphosphate Synthase for Development of Irregular Coupling Activity.
Engineering of a Plant Isoprenyl Diphosphate Synthase for Development of Irregular Coupling Activity.
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DOI：
10.1002/cbic.202100465
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发表时间：
2022-01-05
期刊：
影响因子：
3.2
通讯作者：
Warzecha, Heribert
中科院分区：
文献类型：
--
作者：
Gerasymenko, Iryna;Sheludko, Yuriy, V;Fuertes, Ismael Navarro;Schmidts, Volker;Steinel, Lara;Haumann, Elisabeth;Warzecha, Heribert
我们对一个普通的异戊二磷酸合成酶(IDs)进行了突变，它是茄子番茄中的Neryl二磷酸合成酶(SlNPPS)，它具有两个二甲基烯丙基二磷酸(DMAPP)单元的非头尾偶联的结构相关的类似物，来自中间Lavandula x的Lavandulyl二磷酸合成酶(LiLPPS)。野生型SlNPPS催化异戊烯基二磷酸(IPP)和DMAPP顺式偶联，生成Neryl diPO4。然而，如果该酶只加DMAPP，它能够催化两个DMAPP单元的偶联，合成两个不规则的单萜二磷酸；通过对它们的脱磷产物的核磁共振分析，确定了它们的结构。其中一种酒精是拉凡杜罗。第二个化合物是普拉可可的反式异构体，这是第一个具有这种立体化学构型的不规则环丁烷单萜的例子。SLNPPS的不规则活性占其正常活性的0.4 %，只有在没有IPP的情况下才能显示出该酶的非规律性。天冬酰胺88与组氨酸的交换显著增强了非头尾偶联。在没有IPP的情况下，突变株的非正常活性达到其正常活性的13.1pg/ 。这些结果证明，规则的入侵检测是蛋白质工程的良好起点，目的是开发不规则活性并导致新的单萜结构。规则的异戊二磷酸合成酶的副活性导致两个非头尾萜类结构。对于其中的一种，环丁烷平尾环二磷酸，这是第一个有效的酶合成的例子。合理的蛋白质设计使得不规则活性有相当大的提高。该工程酶可用于昆虫信息素的可持续生产，用于环境友好型害虫防治。
We performed mutagenesis on a regular isoprenyl diphosphate synthase (IDS), neryl diphosphate synthase from Solanum lycopersicum (SlNPPS), that has a structurally related analogue performing non‐head‐to‐tail coupling of two dimethylallyl diphosphate (DMAPP) units, lavandulyl diphosphate synthase from Lavandula x intermedia (LiLPPS). Wild‐type SlNPPS catalyses regular coupling of isopentenyl diphosphate (IPP) and DMAPP in cis‐orientation resulting in the formation of neryl diphosphate. However, if the enzyme is fed with DMAPP only, it is able to catalyse the coupling of two DMAPP units and synthesizes two irregular monoterpene diphosphates; their structures were elucidated by the NMR analysis of their dephosphorylation products. One of the alcohols is lavandulol. The second compound is the trans‐isomer of planococcol, the first example of an irregular cyclobutane monoterpene with this stereochemical configuration. The irregular activity of SlNPPS constitutes 0.4 % of its regular activity and is revealed only if the enzyme is supplied with DMAPP in the absence of IPP. The exchange of asparagine 88 for histidine considerably enhanced the non‐head‐to‐tail coupling. While still only observed in the absence of IPP, irregular activity of the mutant reaches 13.1 % of its regular activity. The obtained results prove that regular IDS are promising starting points for protein engineering aiming at the development of irregular activities and leading to novel monoterpene structures. A side activity of a regular isoprenyl diphosphate synthase results in two non‐head‐to‐tail terpene structures. For one of them, a cyclobutane planococcyl diphosphate, this is the first example of an efficient enzymatic synthesis. Rational design of the protein allowed for a considerable increase of the irregular activity. The engineered enzyme can be implicated in the sustainable production of insect pheromones for environment‐friendly pest control.
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影响因子：
6.1
作者：
Passaro, LC;Webster, FX
通讯作者：
Webster, FX
影响因子：
11.9
作者：
Gao Y;Honzatko RB;Peters RJ
通讯作者：
Peters RJ
影响因子：
4.1
作者：
Chan YT;Ko TP;Yao SH;Chen YW;Lee CC;Wang AH
通讯作者：
Wang AH
影响因子：
16.6
作者：
Teufel, Robin;Kaysser, Leonard;Villaume, Matthew T.;Diethelm, Stefan;Carbullido, Mary K.;Baran, Phil S.;Moore, Bradley S.
通讯作者：
Moore, Bradley S.
DOI：
10.1073/pnas.0904113106
发表时间：
2009-06-30
影响因子：
11.1
作者：
Schilmiller, Anthony L.;Schauvinhold, Ines;Pichersky, Eran
通讯作者：
Pichersky, Eran