Structure of iridoid synthase in complex with NADP+/8-oxogeranial reveals the structural basis of its substrate specificity
Structure of iridoid synthase in complex with NADP+/8-oxogeranial reveals the structural basis of its substrate specificity
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环烯醚萜合酶与 NADP/8-oxogeranial 复合物的结构揭示了其底物特异性的结构基础。
DOI：
10.1016/j.jsb.2016.02.010
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发表时间：
2016-05-01
影响因子：
3
通讯作者：
Zhang, Rongguang
中科院分区：
文献类型：
--
作者：
Qin, Lili;Zhu, Yun;Zhang, Rongguang
环烯醚萜合成酶（Iridoid synthase，IS）是短链脱氢酶/还原酶（Short-chain dehydrogenase/reductase，SDR）超家族中的一种植物酶，可为下游生物碱（包括市售的生物碱、长春碱和长春新碱）提供环状骨架，具有多种药理活性。在这里，我们提出了在脱辅基状态和在与NADP（+）/8-oxogeranial复合的IS的晶体结构，表现出一个活性中心，缺乏经典的酪氨酸/赖氨酸/丝氨酸三联体空间保守的SDRs，只有催化关键功能的三联体酪氨酸保留在Tyr 178。与此一致，Tyrl 78突变为苯丙氨酸残基显著消除了IS的催化活性。在底物结合口袋内，线形的8-oxogeranial采用完全延伸的构象，其两个醛端分别与Tyr 178-OH和Ser 349-OH氢键结合。此外，结合底物的中间碳链被良好有序的疏水支架所包容，包括残基Ile 45，Phe 149，Leu 203，Met 213，Phe 342，I1-345和Leu 352。诱变研究表明，Ser 349和周围的疏水残基是底物特异性的决定因素，因此，催化活性的IS。相反，Gly 150-Pro 160环先前提出作为参与底物结合的因子可能具有非常有限的贡献，因为残基Ile 151-His 161的缺失对催化活性仅具有轻微的影响。我们相信，目前的工作将有助于阐明底物特异性的IS和整合其详细的催化机制。(C)2016 Elsevier Inc. All rights reserved.
Iridoid synthase (IS), as a vegetal enzyme belonging to the short-chain dehydrogenase/reductase (SDR) superfamily, produces the ring skeletons for downstream alkaloids with various pharmaceutical activities, including the commercially available antineoplastic agents, vinblastine and vincristine. Here, we present the crystal structures of IS in apo state and in complex with NADP(+)/8-oxogeranial, exhibiting an active center that lacks the classical Tyr/Lys/Ser triad spatially conserved in SDRs, with only the catalytically critical function of triad tyrosine remained in Tyr178. In consistent, mutation of Tyrl 78 to a phenylalanine residue significantly abolished the catalytic activity of IS. Within the substrate binding pocket, the linear-shaped 8-oxogeranial adopts an entirely extended conformation with its two aldehyde ends hydrogen-bonded to Tyr178-OH and Ser349-OH, respectively. In addition, the intermediate carbon chain of bound substrate is harbored-by a well-ordered hydrophobic scaffold, involving residues Ile45, Phe149, Leu203, Met213, Phe342, I1-345 and Leu352. Mutagenesis studies showed that both Ser349 and the hydrophobic residues around are determinant to the substrate specificity and, consequently, the catalytic activity of IS. In contrast, the Gly150-Pro160 loop previously proposed as a factor involved in substrate binding might have very limited contribution, because the deletion of residues Ile 151-His161 has only slight influence on the catalytic activity. We believe that the present work will help to elucidate the substrate specificity of IS and to integrate its detailed catalytic mechanism. (C) 2016 Elsevier Inc. All rights reserved.