Stereospecificity of Enoylreductase Domains from Modular Polyketide Synthases
Stereospecificity of Enoylreductase Domains from Modular Polyketide Synthases
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模块化聚酮化合物合成酶烯酰还原酶结构域的立体特异性
DOI：
10.1021/acschembio.7b00982
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发表时间：
2018
影响因子：
4
通讯作者：
Jianting Zheng
中科院分区：
文献类型：
--
作者：
Luyun Zhang;Junjie Ji;Meijuan Yuan;Yuanyuan Feng;Lei Wang;Zixin Deng;Linquan Bai;Jianting Zheng
在由酰基载体蛋白(ACP)结构域携带的聚酮化合物中间体的加工过程中，募集甲基丙二酸延伸单元的聚酮化合物合酶模块的烯酰还原酶(ER)结构域将C2甲基分支设置为SorR构型。在本研究中，使用泛硫氨酸和 ACP 结合的反式-2-甲基巴豆酰基底物替代物来检查 ER 结构域的立体特异性。泛硫氨酸结合的硫酯被还原为 2R 和 2S 产物的混合物，而当使用同源 ACP 结合的底物替代物时，几乎完全生成预期的 2Sepimer。 ER 与非同源 ACP 携带的底物替代物的类似孵育显着增加了意想不到的 2Repimer 的产生，突出了立体特异性对 ER 和 ACP 结构域之间适当的蛋白质-蛋白质相互作用的依赖性。 ER 突变体测定揭示了保守的酪氨酸和赖氨酸参与立体控制。总而言之，这些结果扩展了当前对 ER 立体化学的理解，并有助于模块化 PKS 的工程设计。
An enoylreductase (ER) domain of a polyketide synthase module recruiting a methylmalonate extender unit sets the C2 methyl branch to either theSorRconfiguration during processing of a polyketide intermediate carried by an acyl carrier protein (ACP) domain. In the present study, pantetheine- and ACP-boundtrans-2-methylcrotonyl substrate surrogates were used to scrutinize the stereospecificity of the ER domains. The pantetheine-bound thioester was reduced to mixtures of both 2Rand 2Sproducts, whereas the expected 2Sepimer was almost exclusively generated when the cognate ACP-bound substrate surrogate was utilized. The analogous incubation of an ER with the substrate surrogate carried by a noncognate ACP significantly increased the generation of the unexpected 2Repimer, highlighting the dependence of stereospecificity on proper protein–protein interactions between ER and ACP domains. The ER mutant assays revealed the involvement of the conserved tyrosine and lysine in stereocontrol. Taken together, these results expand the current understanding of the ER stereochemistry and help in the engineering of modular PKSs.