Cloned (Comment) | Organism |
---|---|
gene Ldb1010, recombinant expression of His-tagged D2-HDH in Escherichia coli BL21(DE3) pLysS. The enzyme containing the non-native C-terminal hexahistidine tag and a 4-residue linker (Thr Ala Ser Gly linker) is enzymatically active | Lactobacillus delbrueckii subsp. bulgaricus |
Crystallization (Comment) | Organism |
---|---|
purified enzyme in apoform and complexed with coenzyme NAD+, hanging drop vapor diffusion method, mixing of 400 nl of 10 mg/ml protein in 40 mM HEPES, pH 7.4, 300 mM NaCl, and 0.02% v/v monothioglycerol, with 400 nl reservoir solution containing 25% PEG 3350, 200 mM MgCl2, 100 mM HEPES, pH 7.5, and equilibration against 0.1 ml of reservoir solution at 19°C, crystals are supplemented with 10 mM NAD+ for the enzyme complex crystals, X-ray diffraction structure determination at 3.45 A and 2.75 A resolution, respectively, molecular replacement and modeling using the monomer structure of D-2-hydroxyisocaproate dehydrogenase (D-HicDH) from Lactobacillus casei, PDB ID 1DXY | Lactobacillus delbrueckii subsp. bulgaricus |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
additional information | - |
additional information | Michaelis-Menten kinetics, recombinant enzyme | Lactobacillus delbrueckii subsp. bulgaricus | |
0.0591 | - |
phenylpyruvate | recombinant His-tagged enzyme, pH and temperature not specified in the publication | Lactobacillus delbrueckii subsp. bulgaricus |
Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
---|---|---|---|---|---|---|
(R)-2-hydroxycarboxylate + NAD+ | Lactobacillus delbrueckii subsp. bulgaricus | - |
a 2-oxocarboxylate + NADH + H+ | - |
r | |
(R)-2-hydroxycarboxylate + NAD+ | Lactobacillus delbrueckii subsp. bulgaricus ATCC 11842 | - |
a 2-oxocarboxylate + NADH + H+ | - |
r |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Lactobacillus delbrueckii subsp. bulgaricus | Q1GAA2 | - |
- |
Lactobacillus delbrueckii subsp. bulgaricus ATCC 11842 | Q1GAA2 | - |
- |
Purification (Comment) | Organism |
---|---|
recombinant His-tagged D2-HDH from Escherichia coli BL21(DE3) pLysS by nickel affinity chromatography and gel filtration | Lactobacillus delbrueckii subsp. bulgaricus |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
(R)-2-hydroxycarboxylate + NAD+ | - |
Lactobacillus delbrueckii subsp. bulgaricus | a 2-oxocarboxylate + NADH + H+ | - |
r | |
(R)-2-hydroxycarboxylate + NAD+ | - |
Lactobacillus delbrueckii subsp. bulgaricus ATCC 11842 | a 2-oxocarboxylate + NADH + H+ | - |
r | |
phenylpyruvate + NADH + H+ | - |
Lactobacillus delbrueckii subsp. bulgaricus | phenyl-D-lactate + NAD+ | - |
r | |
phenylpyruvate + NADH + H+ | - |
Lactobacillus delbrueckii subsp. bulgaricus ATCC 11842 | phenyl-D-lactate + NAD+ | - |
r |
Subunits | Comment | Organism |
---|---|---|
More | enzyme three-dimensional structure analysis, overview | Lactobacillus delbrueckii subsp. bulgaricus |
Synonyms | Comment | Organism |
---|---|---|
D-isomer specific 2-hydroxyacid dehydrogenase | - |
Lactobacillus delbrueckii subsp. bulgaricus |
D2-HDH | - |
Lactobacillus delbrueckii subsp. bulgaricus |
Ldb1010 | - |
Lactobacillus delbrueckii subsp. bulgaricus |
Turnover Number Minimum [1/s] | Turnover Number Maximum [1/s] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
8.48 | - |
phenylpyruvate | recombinant His-tagged enzyme, pH and temperature not specified in the publication | Lactobacillus delbrueckii subsp. bulgaricus |
Cofactor | Comment | Organism | Structure |
---|---|---|---|
NAD+ | enzyme binding structure analysis, overview | Lactobacillus delbrueckii subsp. bulgaricus | |
NADH | - |
Lactobacillus delbrueckii subsp. bulgaricus |
General Information | Comment | Organism |
---|---|---|
evolution | the enzyme belongs to the the NAD-dependent dehydrogenase family. Comparison with closely related members of the NAD-dependent dehydrogenase family reveals that whilst the D2-HDH core fold is structurally conserved, the substrate-binding site has a number of non-canonical features that may influence substrate selection and thus dictate the physiological function of the enzyme. The protein, 2-hydroxyisocaproate dehydrogenase (HO-HxoDH), is virtually identical to the D2-HDH, with only three amino-acid differences between the two proteins, all at sites not known to be biologically relevant | Lactobacillus delbrueckii subsp. bulgaricus |
additional information | enzyme three-dimensional structure analysis, active site and cofactor binding site structures, overview | Lactobacillus delbrueckii subsp. bulgaricus |
physiological function | the substrate-binding site has a number of non-canonical features that may influence substrate selection and thus dictate the physiological function of the enzyme | Lactobacillus delbrueckii subsp. bulgaricus |