Structural and functional studies reveal the molecular basis of substrate promiscuity of a glycosyltransferase originating from a major agricultural pest.

The two-spotted spider mite, Tetranychus urticae, is a major cosmopolitan pest that feeds on more than 1100 plant species. Its genome contains an unprecedentedly large number of genes involved in detoxifying and transporting xenobiotics, including 80 genes that code for UDP glycosyltransferases (UGTs). These enzymes were acquired via horizontal gene transfer from bacteria after loss in the Chelicerata lineage. UGTs are well-known for their role in phase II metabolism; however, their contribution to host adaptation and acaricide resistance in arthropods, such as T. urticae, is not yet resolved. TuUGT202A2 (Tetur22g00270) has been linked to the ability of this pest to adapt to tomato plants. Moreover, it was shown that this enzyme can glycosylate a wide range of flavonoids. To understand this relationship at the molecular level, structural, functional, and computational studies were performed. Structural studies provided specific snapshots of the enzyme in different catalytically relevant stages. The crystal structure of TuUGT202A2 in complex with UDP-glucose was obtained and site-directed mutagenesis paired with molecular dynamic simulations revealed a novel lid-like mechanism involved in the binding of the activated sugar donor. Two additional TuUGT202A2 crystal complexes, UDP-(S)-naringenin and UDP-naringin, demonstrated that this enzyme has a highly plastic and open-ended acceptor-binding site. Overall, this work reveals the molecular basis of substrate promiscuity of TuUGT202A2 and provides novel insights into the structural mechanism of UGTs catalysis.

二斑叶螨（Tetranychus urticae）是一种主要的世界性害虫，以1100多种植物为食。其基因组包含数量空前多的参与异源物质解毒和转运的基因，其中包括80个编码尿苷二磷酸葡萄糖醛酸转移酶（UGTs）的基因。这些酶是在螯肢动物谱系缺失后通过从细菌水平基因转移获得的。UGTs以其在II相代谢中的作用而闻名；然而，它们对节肢动物（如二斑叶螨）的宿主适应和杀螨剂抗性的贡献尚未明确。TuUGT202A2（Tetur22g00270）与这种害虫适应番茄植株的能力有关。此外，研究表明该酶能够糖基化多种黄酮类化合物。为了在分子水平上理解这种关系，进行了结构、功能和计算研究。结构研究提供了酶在不同催化相关阶段的特定快照。获得了TuUGT202A2与尿苷二磷酸 - 葡萄糖复合物的晶体结构，定点突变结合分子动力学模拟揭示了一种参与活化糖供体结合的新型盖状机制。另外两个TuUGT202A2晶体复合物，尿苷二磷酸 -（S） - 柚皮素和尿苷二磷酸 - 柚皮苷，表明该酶具有高度可塑性和开放式的受体结合位点。总体而言，这项工作揭示了TuUGT202A2底物混杂性的分子基础，并为UGTs催化的结构机制提供了新的见解。