Characterization and Structural Analysis of Emodin-O-Methyltransferase from Aspergillus terreus,Journal of Agricultural and Food Chemistry

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Characterization and Structural Analysis of Emodin-O-Methyltransferase from Aspergillus terreus
Journal of Agricultural and Food Chemistry ( IF 5.7 ) Pub Date : 2022-04-27 , DOI: 10.1021/acs.jafc.2c01281
Yingying Xue ₁ , Yajing Liang _{2,

3,

4} , Wei Zhang _{2,

3,

4,

5} , Ce Geng _{2,

3,

4} , Dandan Feng _{2,

3,

4} , Xuenian Huang _{2,

3,

4,

5} , Sheng Dong _{2,

3,

4} , Yingfang Zhang ₁ , Jia Sun ₁ , Feifei Qi _{2,

3,

4} , Xuefeng Lu _{2,

3,

4,

5,

6}

Affiliation

All O-methylated derivatives of emodin, including physcion, questin, and 1-O-methylemodin, show potential antifungal activities. Notably, emodin and questin are two pivotal intermediates of geodin biosynthesis in Aspergillus terreus. Although most of the geodin biosynthetic steps have been investigated, the key O-methyltransferase (OMT) responsible for the O-methylation of emodin to generate questin has remained unidentified. Herein, through phylogenetic tree analysis and in vitro biochemical assays, the long-sought class II emodin-O-methyltransferase GedA has been functionally characterized. Additionally, the catalytic mechanism and key residues at the catalytic site of GedA were elucidated by enzyme–substrate–methyl donor analogue ternary complex crystal structure determination and site-directed mutagenesis. As we demonstrate, GedA adopts a typical general acid/base (E446/H373)-mediated transmethylation mechanism. In particular, residue D374 is also crucial for efficient catalysis through blocking the formation of intramolecular hydrogen bonds in emodin. This study will facilitate future engineering of GedA for the production of physcion or other site-specific O-methylated anthraquinone derivatives with potential applications as biopesticides.

中文翻译：

土曲霉大黄素-O-甲基转移酶的表征及结构分析

大黄素的所有O-甲基化衍生物，包括 physcion、questin 和 1- O-甲基大黄素，都显示出潜在的抗真菌活性。值得注意的是，大黄素和questin是土曲霉中geodin生物合成的两个关键中间体。尽管已经研究了大多数大地黄素生物合成步骤，但负责大黄素O甲基化生成 questin的关键O甲基转移酶 (OMT) 仍未确定。在此，通过系统发育树分析和体外生化测定，长期寻找的 II 类大黄素-O-甲基转移酶 GedA 已在功能上进行了表征。此外，通过酶 - 底物 - 甲基供体类似物三元复合物晶体结构测定和定点诱变阐明了 GedA 催化位点的催化机制和关键残基。正如我们所证明的，GedA 采用典型的一般酸/碱 (E446/H373) 介导的转甲基化机制。特别是，残基 D374 通过阻断大黄素中分子内氢键的形成对于有效催化也至关重要。这项研究将促进 GedA 的未来工程，用于生产 physcion 或其他位点特异性O-甲基化蒽醌衍生物，并具有作为生物农药的潜在应用。

更新日期：2022-04-27

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