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Literature summary extracted from

  • Furukawa, N.; Miyanaga, A.; Nakajima, M.; Taguchi, H.
    The ternary complex structure of D-mandelate dehydrogenase with NADH and anilino(oxo)acetate (2017), Biochem. Biophys. Res. Commun., 486, 665-670 .
    View publication on PubMed

Cloned(Commentary)

EC Number Cloned (Comment) Organism
1.1.1.379 recombinant enzyme expression in Escherichia coli strain BL21-Gold (DE3) pLysS Enterococcus faecium

Crystallization (Commentary)

EC Number Crystallization (Comment) Organism
1.1.1.379 purified enzyme in complex with substrates NADH and anilino(oxo)acetate (AOA), hanging-drop vapor diffusion method, mixing of 10 mg/ml protein in 5 mM sodium MOPS, pH 7.5, 2 mM NADH, and 20 mM AOA with reservoir solution containing 0.1 M cacodylate-Na, pH 5.0, 0.17 M ammonium sulfate, and 22.5% PEG 8000, in a 1:1 ratio, at 25°C, X-ray diffraction structure determination and analysis at 2.4 A resolution, molecular replacement using the crystal structure of unliganded D-ManDH2 (PDB ID 3WFI) as a search model, modeling. The ternary complex of D-ManDH2 forms a 2fold symmetric homodimer in a crystallographic asymmetric unit, as in the cases of the apo and binary complex structures Enterococcus faecium

Natural Substrates/ Products (Substrates)

EC Number Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
1.1.1.379 phenylglyoxylate + NADH + H+ Enterococcus faecium
-
 (R)-mandelate + NAD+
-
 ?

Organism

EC Number Organism UniProt Comment Textmining
1.1.1.379 Enterococcus faecium
-
-
-

Purification (Commentary)

EC Number Purification (Comment) Organism
1.1.1.379 recombinant enzyme from Escherichia coli strain BL21-Gold (DE3) pLysS by ammonium sulfate fractionation, hydrophobic interaction chromatography, and anion exchange chromatography Enterococcus faecium

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
1.1.1.379 2-oxocaproate + NADH + H+
-
 Enterococcus faecium ? + NAD+
-
 ?
1.1.1.379 2-oxoisovalerate + NADH + H+
-
 Enterococcus faecium ? + NAD+
-
 ?
1.1.1.379 2-oxopantoate + NADH + H+
-
 Enterococcus faecium ? + NAD+
-
 ?
1.1.1.379 anilino(oxo)acetate + NADH + H+ AOA Enterococcus faecium anilino(hydroxy)acetate + NAD+
-
 ?
1.1.1.379 additional information the enzyme exhibits broad substrate specificity toward bulky hydrophobic 2-ketoacids, preferring C3-branched substrates Enterococcus faecium ?
-
-
1.1.1.379 phenylglyoxylate + NADH + H+
-
 Enterococcus faecium (R)-mandelate + NAD+
-
 ?
1.1.1.379 phenylglyoxylate + NADH + H+ i.e. benzoylformate Enterococcus faecium (R)-mandelate + NAD+
-
 ?
1.1.1.379 phenylpyruvate + NADH + H+
-
 Enterococcus faecium ? + NAD+
-
 ?

Subunits

EC Number Subunits Comment Organism
1.1.1.379 homodimer D-ManDH2 constitutively forms a dimeric structure. Each of the two subunits binds the NADH and AOA molecules between the N-terminal (residues 1-176) and C-terminal (residues 180-311). The intersubunit contact is formed between the two C-terminal domains of the dimer on the opposite side of NADH and AOA, and exhibits no significant structural change among the three D-ManDH2 structures. The N-terminal domain exhibits a shear motion along the C-terminal domain between the binary and ternary complex structures, following the domain closure through the hinge motion between the apo and binary complex structures. Residue Asn105 forms hydrogen bonds with both the nicotinamide-ribose of NADH and the carbonyl oxygen of AOA in the ternary complex structure, suggesting that this residue promotes the shear motion of the N-terminal domain Enterococcus faecium

Synonyms

EC Number Synonyms Comment Organism
1.1.1.379 D-mandelate dehydrogenase
-
 Enterococcus faecium
1.1.1.379 D-ManDH
-
 Enterococcus faecium
1.1.1.379 D-ManDH2
-
 Enterococcus faecium
1.1.1.379 NAD-dependent D-mandelate dehydrogenase
-
 Enterococcus faecium

Temperature Optimum [°C]

EC Number Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
1.1.1.379 30
-
 assay at Enterococcus faecium

pH Optimum

EC Number pH Optimum Minimum pH Optimum Maximum Comment Organism
1.1.1.379 additional information
-
 D-ManDH2 only shows gradually decreasing Vmax/Km values in the 2-oxoacid reduction with increasing pH, without a marked change in substrate preference Enterococcus faecium
1.1.1.379 4.5 7 dependent on substrate, overview Enterococcus faecium

Cofactor

EC Number Cofactor Comment Organism Structure
1.1.1.379 NADH the N-terminal domain forms many more hydrogen bonds with NADH than the C-terminal domain in the binary complex, and moves together with the bound NADH molecule in the shear motion, these intermolecular hydrogen bonds are retained Enterococcus faecium

General Information

EC Number General Information Comment Organism
1.1.1.379 evolution the enzyme belongs to a ketopantoate reductase (KPR)-related D-2-hydroxyacid dehydrogenase family Enterococcus faecium
1.1.1.379 additional information enzyme D-ManDH2 forms many hydrogen bonds with the 2-ketoacid moiety of AOA in the ternary complex structure, involving Asn105, Lys187, Asn191, Asn195 and Ser260, substrate binding structures in binary and tertiary complexes, structure and structure-function analysis, overview. The substrate binding induces a shear motion of the N-terminal domain along the C-terminal domain, following the hinge motion induced by the NADH binding, and allows the bound NADH molecule to form favorable interactions with a 2-oxoacid substrate. D-ManDH possesses a sufficiently wide pocket that accommodates the C3 branched side chains of substrates like KPR, but unlike the pocket of KPR, the pocket of D-ManDH comprises an entirely hydrophobic surface and an expanded space, in which the AOA benzene is accommodated. The expanded space mostly comprises a mobile loop structure, which likely modulates the shape and size of the space depending on the substrate. D-ManDH2 undergoes the opening and closing motion in the entrance area of the substrate binding pocket, which comprises mostly Met128, Thr130 and Ile254, between the binary and ternary complexes. Lys187 is actually the acid/base catalyst of the enzyme Enterococcus faecium