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Stereospecificity of Enoylreductase Domains from Modular Polyketide Synthases
ACS Chemical Biology ( IF 4 ) Pub Date : 2018-02-13 00:00:00 , DOI: 10.1021/acschembio.7b00982 Luyun Zhang 1 , Junjie Ji 2 , Meijuan Yuan 1 , Yuanyuan Feng 1 , Lei Wang 1 , Zixin Deng 1 , Linquan Bai 1 , Jianting Zheng 1
ACS Chemical Biology ( IF 4 ) Pub Date : 2018-02-13 00:00:00 , DOI: 10.1021/acschembio.7b00982 Luyun Zhang 1 , Junjie Ji 2 , Meijuan Yuan 1 , Yuanyuan Feng 1 , Lei Wang 1 , Zixin Deng 1 , Linquan Bai 1 , Jianting Zheng 1
Affiliation
An enoylreductase (ER) domain of a polyketide synthase module recruiting a methylmalonate extender unit sets the C2 methyl branch to either the S or R configuration during processing of a polyketide intermediate carried by an acyl carrier protein (ACP) domain. In the present study, pantetheine- and ACP-bound trans-2-methylcrotonyl substrate surrogates were used to scrutinize the stereospecificity of the ER domains. The pantetheine-bound thioester was reduced to mixtures of both 2R and 2S products, whereas the expected 2S epimer 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 2R epimer, 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.
中文翻译:
模块化聚酮合酶的烯丙基还原酶结构域的立体特异性。
募集甲基丙二酸扩展剂单元的聚酮化合物合酶模块的烯基还原酶(ER)域在酰基载体蛋白(ACP)域携带的聚酮化合物中间体的加工过程中将C2甲基分支设置为S或R构型。在本研究中,泛胺结合和ACP绑定反-2-甲基巴豆酰基底物替代物被用来审查ER域的立体特异性。泛胺键合的硫酯还原为2 R和2 S产物的混合物,而预期的2 S当使用同源的ACP结合的底物替代物时,差向异构体几乎全部生成。ER与非同源ACP携带的底物替代物的类似孵育,显着增加了意外2 R差向异构体的生成,突显了立体特异性对ER与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了立体控制。综上所述,这些结果扩大了对ER立体化学的当前理解,并有助于模块化PKS的工程设计。
更新日期:2018-02-13
中文翻译:
模块化聚酮合酶的烯丙基还原酶结构域的立体特异性。
募集甲基丙二酸扩展剂单元的聚酮化合物合酶模块的烯基还原酶(ER)域在酰基载体蛋白(ACP)域携带的聚酮化合物中间体的加工过程中将C2甲基分支设置为S或R构型。在本研究中,泛胺结合和ACP绑定反-2-甲基巴豆酰基底物替代物被用来审查ER域的立体特异性。泛胺键合的硫酯还原为2 R和2 S产物的混合物,而预期的2 S当使用同源的ACP结合的底物替代物时,差向异构体几乎全部生成。ER与非同源ACP携带的底物替代物的类似孵育,显着增加了意外2 R差向异构体的生成,突显了立体特异性对ER与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了立体控制。综上所述,这些结果扩大了对ER立体化学的当前理解,并有助于模块化PKS的工程设计。