Molecular Insights into the Regioselectivity of the Fe(II)/2-Ketoglutarate-Dependent Dioxygenase-Catalyzed C–H Hydroxylation of Amino Acids
Molecular Insights into the Regioselectivity of the Fe(II)/2-Ketoglutarate-Dependent Dioxygenase-Catalyzed C–H Hydroxylation of Amino Acids
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Fe(II)/2-酮戊二酸依赖性双加氧酶催化的氨基酸 C-H 羟基化区域选择性的分子洞察
DOI：
10.1021/acscatal.2c03106
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发表时间：
2022-09
期刊：
影响因子：
12.9
通讯作者：
Yao Nie
中科院分区：
文献类型：
--
作者：
Lunjie Wu;Jianhong An;Xiaoran Jing;Chun-Chi Chen;Longhai Dai;Yan Xu;Weidong Liu;Rey-Ting Guo;Yao Nie
L-异亮氨酸双加氧酶 (IDO) 直接催化几种疏水性脂肪族氨基酸的 C-H 键羟基化。然而，IDO 的不明确选择性阻碍了其在手性羟基氨基酸生产中的应用。利用IDO对l-正亮氨酸进行羟基化，生成具有明显区域选择性的4-羟基正亮氨酸和5-羟基正亮氨酸，研究了IDO区域选择性的机制。结合计算结构分析和高通量筛选，IDO 结构揭示了具有增强区域选择性的单位点变体（T244A、T244G 和 T244S）；例如，区域异构体产品中的 4-羟基正亮氨酸纯度从 78.9%（通过野生型 IDO）分别提高到 95.1%、96.6% 和 95.3%。分子动力学模拟表明，将 T244 突变为更小的氨基酸可微调底物结合姿势。对于需要精确定位的不对称催化，这种变化扩大了底物 C4 或 C5 与 Fe2+ 之间最常见的距离，使 4-羟基正亮氨酸的最大纯度达到 96.6%。我们提高了对 Fe(II)/2-酮戊二酸依赖性双加氧酶的区域选择性的理解，并为基于 C-H 羟基化的活性化合物多样化提供了一条途径。
l-Isoleucine dioxygenase (IDO) directly catalyzes the C–H bond hydroxylation of several hydrophobic aliphatic amino acids. However, the ambiguous selectivity of IDO prevents its application in chiral hydroxy amino acid production. The hydroxylation ofl-norleucine by IDO, which produces 4-hydroxynorleucine and 5-hydroxynorleucine with obvious regioselectivity, was used to investigate the mechanism of IDO regioselectivity. Along with computational structural analysis and high-throughput screening, the IDO structure revealed single-site variants (T244A, T244G, and T244S) with enhanced regioselectivity; for example, the 4-hydroxynorleucine purity in regioisomeric products increased from 78.9% (by wild-type IDO) to 95.1%, 96.6%, and 95.3%, respectively. Molecular dynamics simulations showed that mutating T244 into smaller amino acids fine-tuned the substrate binding pose. For asymmetric catalysis requiring precise positioning, this change expanded the most frequent distances between the substrate C4 or C5 and Fe2+, giving a maximum 4-hydroxynorleucine purity of 96.6%. We improved the understanding of the regioselectivity of Fe(II)/2-ketoglutarate-dependent dioxygenases and provide a route for diversifying C–H hydroxylation-based active compounds.
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影响因子：
2.9
作者：
Xu, Qingping;Grant, Joanna;Chiu, Hsiu-Ju;Farr, Carol L.;Jaroszewski, Lukasz;Knuth, Mark W.;Miller, Mitchell D.;Lesley, Scott A.;Godzik, Adam;Elsliger, Marc-Andre;Deacon, Ashley M.;Wilson, Ian A.
通讯作者：
Wilson, Ian A.
影响因子：
4.1
作者：
Lawrence, CC;Sobey, WJ;Schofield, CJ
通讯作者：
Schofield, CJ
影响因子：
5.5
作者：
Stroet, Martin;Caron, Bertrand;Mark, Alan E.
通讯作者：
Mark, Alan E.
影响因子：
8.6
作者：
A. Amatuni;A. Shuster;A. Adibekian;Hans Renata
通讯作者：
A. Amatuni;A. Shuster;A. Adibekian;Hans Renata
影响因子：
3.9
作者：
J. Rolf;Philipp Nerke;Annette Britner;S. Krick;S. Lütz;Katrin Rosenthal
通讯作者：
J. Rolf;Philipp Nerke;Annette Britner;S. Krick;S. Lütz;Katrin Rosenthal