Structural basis for olivetolic acid formation by a polyketide cyclase from Cannabis sativa
Structural basis for olivetolic acid formation by a polyketide cyclase from Cannabis sativa
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DOI：
10.1111/febs.13654
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发表时间：
2016-03
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通讯作者：
Xinmei Yang;T. Matsui;T. Kodama;Takahiro Mori;Xiaoxi Zhou;F. Taura;H. Noguchi;I. Abe;H. Morita
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文献类型：
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作者：
Xinmei Yang;T. Matsui;T. Kodama;Takahiro Mori;Xiaoxi Zhou;F. Taura;H. Noguchi;I. Abe;H. Morita
在聚酮生物合成中，环的形成是关键的多样化步骤之一。来自大麻的橄榄酸环化酶（OAC）是目前已知唯一的植物多酮环化酶，参与大麻素的生物合成。此外，它是唯一一个功能表征的植物α+β桶（DABB）蛋白，它催化线性戊基四β酮CoA的C2-C7醛环化，作为底物，生成橄榄醇酸（OA）。本文分别在1.32和1.70 Å分辨率下求解了OAC apo和OAC - oa复合二元晶体结构。晶体结构表明，该酶确实属于DABB超家族，并且具有独特的活性位点空腔，其中包含戊基结合疏水口袋和聚酮结合位点，这在功能和结构上表征的细菌聚酮环化酶中从未观察到过。此外，位点定向诱变研究表明，Tyr72和His78在催化中心起酸/碱催化剂的作用。结构和/或功能研究表明，OAC酶缺乏硫酯酶和芳香酶活性。这些观察结果表明，OAC采用独特的催化机制，利用酸/碱催化化学形成OA前体。因此，在这项工作中获得的结构和功能见解为将来发现的植物聚酮环化酶的分析提供了基础。
In polyketide biosynthesis, ring formation is one of the key diversification steps. Olivetolic acid cyclase (OAC) from Cannabis sativa, involved in cannabinoid biosynthesis, is the only known plant polyketide cyclase. In addition, it is the only functionally characterized plant α+β barrel (DABB) protein that catalyzes the C2–C7 aldol cyclization of the linear pentyl tetra‐β‐ketide CoA as the substrate, to generate olivetolic acid (OA). Herein, we solved the OAC apo and OAC–OA complex binary crystal structures at 1.32 and 1.70 Å resolutions, respectively. The crystal structures revealed that the enzyme indeed belongs to the DABB superfamily, as previously proposed, and possesses a unique active‐site cavity containing the pentyl‐binding hydrophobic pocket and the polyketide binding site, which have never been observed among the functionally and structurally characterized bacterial polyketide cyclases. Furthermore, site‐directed mutagenesis studies indicated that Tyr72 and His78 function as acid/base catalysts at the catalytic center. Structural and/or functional studies of OAC suggested that the enzyme lacks thioesterase and aromatase activities. These observations demonstrated that OAC employs unique catalytic machinery utilizing acid/base catalytic chemistry for the formation of the precursor of OA. The structural and functional insights obtained in this work thus provide the foundation for analyses of the plant polyketide cyclases that will be discovered in the future.