7CLZ

Crystal structure of Alp1U W187F/Y247F in complex with fluostatin C

7CLZ の概要

• エントリーDOI: 10.2210/pdb7clz/pdb
• 関連するPDBエントリー: 6KXH
• 分子名称: Putative hydrolase, Fluostatin C, SODIUM ION, ... (6 entities in total)
• 機能のキーワード: substrate, complex, biosynthesis, hydrolase
• 由来する生物種: Streptomyces ambofaciens (strain ATCC 23877 / 3486 / DSM 40053 / JCM 4204 / NBRC 12836 / NRRL B-2516)
• タンパク質・核酸の鎖数: 4
• 化学式量合計: 149662.36

構造登録者

Zhang, L.,De, B.C. (登録日: 2020-07-22, 公開日: 2020-10-07, 最終更新日: 2023-11-29)

主引用文献

Zhang, L.,De, B.C.,Zhang, W.,Mandi, A.,Fang, Z.,Yang, C.,Zhu, Y.,Kurtan, T.,Zhang, C.
Mutation of an atypical oxirane oxyanion hole improves regioselectivity of the alpha / beta-fold epoxide hydrolase Alp1U.
J.Biol.Chem., 295:16987-16997, 2020

PubMed Abstract: Epoxide hydrolases (EHs) have been characterized and engineered as biocatalysts that convert epoxides to valuable chiral vicinal diol precursors of drugs and bioactive compounds. Nonetheless, the regioselectivity control of the epoxide ring opening by EHs remains challenging. Alp1U is an α/β-fold EH that exhibits poor regioselectivity in the epoxide hydrolysis of fluostatin C (compound 1) and produces a pair of stereoisomers. Herein, we established the absolute configuration of the two stereoisomeric products and determined the crystal structure of Alp1U. A Trp-186/Trp-187/Tyr-247 oxirane oxygen hole was identified in Alp1U that replaced the canonical Tyr/Tyr pair in α/β-EHs. Mutation of residues in the atypical oxirane oxygen hole of Alp1U improved the regioselectivity for epoxide hydrolysis on 1. The single site Y247F mutation led to highly regioselective (98%) attack at C-3 of 1, whereas the double mutation W187F/Y247F resulted in regioselective (94%) nucleophilic attack at C-2. Furthermore, single-crystal X-ray structures of the two regioselective Alp1U variants in complex with 1 were determined. These findings allowed insights into the reaction details of Alp1U and provided a new approach for engineering regioselective epoxide hydrolases.

PubMed: 33004437
DOI: 10.1074/jbc.RA120.015563

実験手法

X-RAY DIFFRACTION (2.10001009432 Å)