4PB4

D-threo-3-hydroxyaspartate dehydratase H351A mutant complexed with 2-amino maleic acid

4PB4 の概要

• エントリーDOI: 10.2210/pdb4pb4/pdb
• 関連するPDBエントリー: 3WQC 3WQD 3WQE 3WQF 3WQG 4PB3 4PB5
• 分子名称: D-threo-3-hydroxyaspartate dehydratase, PYRIDOXAL-5'-PHOSPHATE, MAGNESIUM ION, ... (5 entities in total)
• 機能のキーワード: plp enzyme, dehydratase, metalloprotein, lyase
• 由来する生物種: Delftia sp.
• タンパク質・核酸の鎖数: 2
• 化学式量合計: 83769.16

構造登録者

Yasutake, Y.,Matsumoto, Y.,Wada, M. (登録日: 2014-04-11, 公開日: 2015-03-11, 最終更新日: 2023-12-27)

主引用文献

Matsumoto, Y.,Yasutake, Y.,Takeda, Y.,Tamura, T.,Yokota, A.,Wada, M.
Structural insights into the substrate stereospecificity of D-threo-3-hydroxyaspartate dehydratase from Delftia sp. HT23: a useful enzyme for the synthesis of optically pure L-threo- and D-erythro-3-hydroxyaspartate.
Appl.Microbiol.Biotechnol., 99:7137-7150, 2015

PubMed Abstract: D-threo-3-Hydroxyaspartate dehydratase (D-THA DH) is a fold-type III pyridoxal 5'-phosphate-dependent enzyme, isolated from a soil bacterium of Delftia sp. HT23. It catalyzes the dehydration of D-threo-3-hydroxyaspartate (D-THA) and L-erythro-3-hydroxyaspartate (L-EHA). To elucidate the mechanism of substrate stereospecificity, crystal structures of D-THA DH were determined in complex with various ligands, such as an inhibitor (D-erythro-3-hydroxyaspartate (D-EHA)), a substrate (L-EHA), and the reaction intermediate (2-amino maleic acid). The C (β) -OH of L-EHA occupied a position close to the active-site Mg(2+), clearly indicating a possibility of metal-assisted C (β) -OH elimination from the substrate. In contrast, the C (β) -OH of an inhibitor was bound far from the active-site Mg(2+). This suggests that the substrate specificity of D-THA DH is determined by the orientation of the C (β) -OH at the active site, whose spatial arrangement is compatible with the 3R configuration of 3-hydroxyaspartate. We also report an optically pure synthesis of L-threo-3-hydroxyaspartate (L-THA) and D-EHA, promising intermediates for the synthesis of β-benzyloxyaspartate, by using a purified D-THA DH as a biocatalyst for the resolution of racemic DL-threo-3-hydroxyaspartate (DL-THA) and DL-erythro-3-hydroxyaspartate (DL-EHA). Considering 50 % of the theoretical maximum, efficient yields of L-THA (38.9 %) and D-EHA (48.9 %) as isolated crystals were achieved with >99 % enantiomeric excess (e.e.). The results of nuclear magnetic resonance signals verified the chemical purity of the products. We were directly able to isolate analytically pure compounds by the recrystallization of acidified reaction mixtures (pH 2.0) and thus avoiding the use of environmentally harmful organic solvents for the chromatographic purification.

PubMed: 25715785
DOI: 10.1007/s00253-015-6479-3

実験手法

X-RAY DIFFRACTION (1.8 Å)