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D-Threonine aldolase from the green alga Chlamydomonas reinhardtii (CrDTA) catalyzes the interconversion of several β-hydroxy-D-amino acids (e.g. D-threonine) and glycine plus the corresponding aldehydes. Recombinant CrDTA was overexpressed in Escherichia coli and purified to homogeneity; it was subsequently crystallized using the hanging-drop vapour-diffusion method at 295 K. Data were collected and processed at 1.85 Å resolution. Analysis of the diffraction pattern showed that the crystal belonged to space group P1, with unit-cell parameters a = 64.79, b = 74.10, c = 89.94 Å, α = 77.07, β = 69.34, γ = 71.93°. The asymmetric unit contained four molecules of CrDTA. The Matthews coefficient was calculated to be 2.12 Å Da and the solvent content was 41.9%.

D-Amino-acid oxidases (DAAOs) catalyze the oxidative deamination of neutral and basic D-amino acids. The DAAO from the thermophilic fungus Rasamsonia emersonii strain YA (ReDAAO) has a high thermal stability and a unique broad substrate specificity that includes the acidic D-amino acid D-Glu as well as various neutral and basic D-amino acids. In this study, ReDAAO was crystallized by the hanging-drop vapor-diffusion method and its crystal structure was determined at a resolution of 2.00 Å. The crystal structure of the enzyme revealed that unlike other DAAOs, ReDAAO forms a homotetramer and contains an intramolecular disulfide bond (Cys230–Cys285), suggesting that this disulfide bond is involved in the higher thermal stability of ReDAAO. Moreover, the structure of the active site and its vicinity in ReDAAO indicates that Arg97, Lys99, Lys114 and Ser231 are candidates for recognizing the side chain of D-Glu.

D-Threonine aldolase (DTA) is a pyridoxal-5′-phosphate-dependent enzyme which catalyzes the reversible aldol reaction of glycine with a corresponding aldehyde to yield the D-form β-hydroxy-α-amino acid. This study produced and investigated the crystal structure of DTA from Chlamydomonas reinhardtii (CrDTA) at 1.85 Å resolution. To our knowledge, this is the first report on the crystal structure of eukaryotic DTA. Compared with the structure of bacterial DTA, CrDTA has a similar arrangement of active-site residues. On the other hand, we speculated that some non-conserved residues alter the affinity for substrates and inhibitors. The structure of CrDTA could provide insights into the structural framework for structure-guided protein engineering studies to modify reaction selectivity.

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