4PB5

D-threo-3-hydroxyaspartate dehydratase H351A mutant complexed with L-erythro-3-hydroxyaspartate

Summary for 4PB5

• Entry DOI: 10.2210/pdb4pb5/pdb
• Related: 3WQC 3WQD 3WQE 3WQF 3WQG 4PB3 4PB4
• Descriptor: D-threo-3-hydroxyaspartate dehydratase, PYRIDOXAL-5'-PHOSPHATE, MAGNESIUM ION, ... (6 entities in total)
• Functional Keywords: plp enzyme, dehydratase, metalloprotein, lyase
• Biological source: Delftia sp.
• Total number of polymer chains: 2
• Total formula weight: 83897.29

Authors

Yasutake, Y.,Matsumoto, Y.,Wada, M. (deposition date: 2014-04-11, release date: 2015-03-11, Last modification date: 2023-12-27)

Primary citation

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

Experimental method

X-RAY DIFFRACTION (1.9 Å)