Structure of epi-isozizaene synthase from Streptomyces coelicolor A3(2), a platform for new terpenoid cyclization templates.
Structure of epi-isozizaene synthase from Streptomyces coelicolor A3(2), a platform for new terpenoid cyclization templates.
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DOI：
10.1021/bi902088z
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发表时间：
2010-03-02
期刊：
影响因子：
2.9
通讯作者：
Christianson, David W.
中科院分区：
文献类型：
--
作者：
Aaron, Julie A.;Lin, Xin;Cane, David E.;Christianson, David W.
天蓝色链霉菌A3(2)的重组表异氮烯合成酶(EIZS)是一种倍半萜环化酶，其X-射线晶体结构的分辨率为1.60？具体来说，野生型EIZS的结构是它与3个镁离子、无机焦磷酸盐(PPI)和苄基三乙基阳离子(BTAC)形成的络合物的封闭构象。此外，D99N EIZS的结构已经确定为开放的、无配体的构象，分辨率为1.90？这两种结构的比较提供了细菌萜类环酶底物结合和催化所需的构象变化的第一个视角。此外，BTAC的结合作用可能与碳阳离子中间体在催化中的作用类似。因此，F95、F96和F198的芳环通过阳离子-π相互作用表现出良好的定向，以稳定环化级联中的碳正离子中间体。酶活性部位芳香族残基的突变导致碳正离子中间体稳定模式的改变以及法尼基二磷酸环化模板的改变而导致替代倍半萜产物阵列的产生。相应地，F198A EIZS与3个镁离子、PPI和BTAC络合的1.64？分辨率的晶体结构揭示了BTAC的另一种结合方向；碳正离子中间体的另一种结合方向可能导致替代产物的形成。最后，测定了4个Hg2+离子与野生型EIZS的晶体结构，结果表明，金属结合引起了活性中心G螺旋的显著构象变化。
The X-ray crystal structure of recombinant epi-isozizaene synthase (EIZS), a sesquiterpene cyclase from Streptomyces coelicolor A3(2), has been determined at 1.60 Å resolution. Specifically, the structure of wild-type EIZS is that of its closed conformation in complex with 3 Mg2+ ions, inorganic pyrophosphate (PPi), and the benzyltriethylammonium cation (BTAC). Additionally, the structure of D99N EIZS has been determined in an open, ligand-free conformation at 1.90 Å resolution. Comparison of these two structures provides the first view of conformational changes required for substrate binding and catalysis in a bacterial terpenoid cyclase. Moreover, the binding interactions of BTAC may mimic those of a carbocation intermediate in catalysis. Accordingly, the aromatic rings of F95, F96, and F198 appear well-oriented to stabilize carbocation intermediates in the cyclization cascade through cation-π interactions. Mutagenesis of aromatic residues in the enzyme active site results in the production of alternative sesquiterpene product arrays due to altered modes of stabilization of carbocation intermediates as well as altered templates for the cyclization of farnesyl diphosphate. Accordingly, the 1.64 Å resolution crystal structure of F198A EIZS complexed with 3 Mg2+ ions, PPi, and BTAC reveals an alternative binding orientation of BTAC; alternative binding orientations of a carbocation intermediate could lead to the formation of alternative products. Finally, the crystal structure of wild-type EIZS complexed with 4 Hg2+ ions has been determined at 1.90 Å resolution, showing that metal binding triggers a significant conformational change of helix G to cap the active site.
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DOI：
10.1107/s0907444904019158
发表时间：
2004-12-01
影响因子：
2.2
作者：
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发表时间：
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影响因子：
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