Crystal structure and functional analysis of tetracenomycin ARO/CYC: Implications for cyclization specificity of aromatic polyketides
Crystal structure and functional analysis of tetracenomycin ARO/CYC: Implications for cyclization specificity of aromatic polyketides
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DOI：
10.1073/pnas.0709223105
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发表时间：
2008-04-08
影响因子：
11.1
通讯作者：
Tsai, Shiou-Chuan
中科院分区：
文献类型：
--
作者：
Ames, Brian Douglas;Korman, Tyler Paz;Tsai, Shiou-Chuan
聚酮化合物是一类具有高度多样化化学结构和药物活性的天然产物。由芳香酶/环化酶 (ARO/CYC) 促进的聚酮化合物环化有助于芳香族聚酮化合物的多样化。由于缺乏第一环 ARO/CYC 结构，ARO/CYC 如何促进高度特异性的环化尚不清楚。 Tcm ARO/CYC 的 1.9 埃晶体结构表明该酶属于具有螺旋握把折叠的 Bet v1 样超家族（或 STAR 结构域家族），并含有高度保守的内部口袋。对接、诱变和体内测定表明，口袋的大小、形状和组成对于 C9-C14 第一环和 C7-C16 第二环环化的聚酮化合物链的定向和特异性折叠非常重要。发现两个口袋残基 R69 和 Y35 对于促进第一环和第二环环化特异性至关重要。不同的口袋残基突变影响聚酮产物的分布。基于结构突变时钟结果提出了一种机制。这些结果强烈表明前两个环的区域特异性环化和随后的芳构化发生在内部口袋中。从这项工作中获得的化学见解为定义 ARO/CYC 环化特异性的分子规则奠定了基础，其合理控制对于未来新型抗癌和抗生素芳香族聚酮化合物的工程生物合成至关重要。
Polyketides are a class of natural products with highly diverse chemical structures and pharmaceutical activities. Polyketide cyclization, promoted by the aromatase/cyclase (ARO/CYC), helps diversify aromatic polyketides. How the ARO/CYC promotes highly specific cyclization is not well understood because of the lack of a first-ring ARO/CYC structure. The 1.9 angstrom crystal structure of Tcm ARO/CYC reveals that the enzyme belongs to the Bet v1-like superfamily (or STAR domain family) with a helix-grip fold, and contains a highly conserved interior pocket. Docking, mutagenesis, and an in vivo assay show that the size, shape, and composition of the pocket are important to orient and specifically fold the polyketide chain for C9-C14 first-ring and C7-C16 second-ring cyclizations. Two pocket residues, R69 and Y35, were found to be essential for promoting first- and second-ring cyclization specificity. Different pocket residue mutations affected the polyketide product distribution. A mechanism is proposed based on the structure-mutation-clocking results. These results strongly suggest that the regiospecific cyclizations of the first two rings and subsequent aromatizations take place in the interior pocket. The chemical insights gleaned from this work pave the foundation toward defining the molecular rules for the ARO/CYC cyclization specificity, whose rational control will be important for future endeavors in the engineered biosynthesis of novel anticancer and antibiotic aromatic polyketides.