Dehydroquinate dehydratase (DHQD) catalyzes the conversion of 3-dehydroquinic acid (DHQ) into 3-dehydroshikimic acid in the mid stage of the shikimate pathway, which is essential for the biosynthesis of aromatic amino acids and folates. Here, we report two the crystal structures of type II DHQD (CgDHQD) derived from Corynebacterium glutamicum, which is a widely used industrial platform organism. We determined the structures for CgDHQD^WT with the citrate at a resolution of 1.80Å and CgDHQD^R19A with DHQ complexed forms at a resolution of 2.00 Å, respectively. The enzyme forms a homododecamer consisting of four trimers with three interfacial active sites. We identified the DHQ-binding site of CgDHQD and observed an unusual binding mode of citrate inhibitor in the site with a half-opened lid loop. A structural comparison of CgDHQD with a homolog derived from Streptomyces coelicolor revealed differences in the terminal regions, lid loop, and active site. Particularly, CgDHQD, including some Corynebacterium species, possesses a distinctive residue P105, which is not conserved in other DHQDs at the position near the 5-hydroxyl group of DHQ. Replacements of P105 with isoleucine and valine, conserved in other DHQDs, caused an approximately 70% decrease in the activity, but replacement of S103 with threonine (CgDHQD^S103T) caused a 10% increase in the activity. Our biochemical studies revealed the importance of key residues and enzyme kinetics for wild type and CgDHQD^S103T, explaining the effect of the variation. This structural and biochemical study provides valuable information for understanding the reaction efficiency that varies due to structural differences caused by the unique sequences of CgDHQD.

中文翻译：

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谷氨酸棒杆菌 3-脱氢奎宁酸脱水酶的结构和生化分析。

脱氢奎尼酸脱水酶 (DHQD) 在莽草酸途径的中期催化 3-脱氢奎尼酸 (DHQ) 转化为 3-脱氢莽草酸，这对于芳香族氨基酸和叶酸的生物合成至关重要。在这里，我们报告了两种源自谷氨酸棒杆菌的 II 型 DHQD ( Cg DHQD)的晶体结构，谷氨酸棒杆菌是一种广泛使用的工业平台生物。我们分别以 1.80 Å 的分辨率确定了含有柠檬酸盐的Cg DHQD ^WT的结构，以 2.00 Å 的分辨率确定了含有 DHQ 复合形式的Cg DHQD ^{R19A的结构。}该酶形成由具有三个界面活性位点的四个三聚体组成的同十二聚体。我们鉴定了Cg DHQD的 DHQ 结合位点，并在半开盖环位点观察到柠檬酸抑制剂的不寻常结合模式。Cg DHQD 与天蓝色链霉菌同源物的结构比较揭示了末端区域、盖环和活性位点的差异。特别地，Cg DHQD，包括一些棒状杆菌属物种，在靠近DHQ的5-羟基的位置处具有在其他DHQD中不保守的独特残基P105。用在其他 DHQD 中保守的异亮氨酸和缬氨酸替换 P105，导致活性降低约 70%，但用苏氨酸 ( Cg DHQD ^S103T )替换 S103导致活性增加 10%。我们的生化研究揭示了野生型和Cg DHQD ^S103T的关键残基和酶动力学的重要性，解释了变异的影响。这项结构和生化研究为理解由于Cg DHQD 独特序列引起的结构差异而变化的反应效率提供了有价值的信息。