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Literature summary for 1.1.1.28 extracted from

  • Zheng, Z.; Sheng, B.; Gao, C.; Zhang, H.; Qin, T.; Ma, C.; Xu, P.
    Highly stereoselective biosynthesis of (R)-alpha-hydroxy carboxylic acids through rationally re-designed mutation of D-lactate dehydrogenase (2013), Sci. Rep., 3, 3401.
    View publication on PubMedView publication on EuropePMC
Search for more literature for 1.1.1.28

Cloned(Commentary)

Cloned (Comment) Organism
gene ldhD, recombinant expression of His-tagged wild-type and mutant enzymes in Escherichia coli strain BL21(DE3) Lactobacillus delbrueckii subsp. bulgaricus

Protein Variants

Protein Variants Comment Organism
F299Y site-directed mutagenesis Lactobacillus delbrueckii subsp. bulgaricus
additional information highly stereoselective biosynthesis of (R)-alpha-hydroxy carboxylic acids through rationally re-designed mutation of D-lactate dehydrogenase, asymmetric reduction of a homologous series of alpha-keto carboxylic acids such as phenylpyruvic acid, 2-oxobutyric acid, 2-oxovaleric acid, beta-hydroxypyruvate, overview. Compared with wild-type D-nLDH, the Y52L mutant D-nLDH shows elevated activities toward unnatural substrates especially with large substitutes at C-3. By the biocatalysis combined with a formate dehydrogenase for in situ generation of NADH, the corresponding (R)-alpha-hydroxy carboxylic acids can be produced at high yields and highly optical purity. Production of chiral (R)-phenyllactic acid. 50 mM PPA is completely reduced to (R)-phenyllactate in 90 min with a high yield of 99.0% and a highly optical purity (99.9% e.e.) by the engineered coupled production system. Activties of the F299Y mutant are similar to the wild-type enzyme Lactobacillus delbrueckii subsp. bulgaricus
Y52L site-directed mutagenesis Lactobacillus delbrueckii subsp. bulgaricus
Y52L/F299Y site-directed mutagenesis Lactobacillus delbrueckii subsp. bulgaricus

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.27
-
 phenylpyruvate pH 7.4, 37°C, recombinant His-taged mutant Y52L Lactobacillus delbrueckii subsp. bulgaricus
0.32
-
 phenylpyruvate pH 7.4, 37°C, recombinant His-taged mutant F299Y Lactobacillus delbrueckii subsp. bulgaricus
1.4
-
 phenylpyruvate pH 7.4, 37°C, recombinant His-taged mutant Y52L/F299Y Lactobacillus delbrueckii subsp. bulgaricus
11.4
-
 phenylpyruvate pH 7.4, 37°C, recombinant His-taged wild-type enzyme Lactobacillus delbrueckii subsp. bulgaricus

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
(R)-lactate + NAD+ Lactobacillus delbrueckii subsp. bulgaricus
-
 pyruvate + NADH + H+
-
 r
(R)-lactate + NAD+ Lactobacillus delbrueckii subsp. bulgaricus ATCC 11842
-
 pyruvate + NADH + H+
-
 r
pyruvate + NADH + H+ Lactobacillus delbrueckii subsp. bulgaricus
-
 (R)-lactate + NAD+
-
 r
pyruvate + NADH + H+ Lactobacillus delbrueckii subsp. bulgaricus ATCC 11842
-
 (R)-lactate + NAD+
-
 r

Organism

Organism UniProt Comment Textmining
Lactobacillus delbrueckii subsp. bulgaricus
-
-
-
Lactobacillus delbrueckii subsp. bulgaricus ATCC 11842
-
-
-

Purification (Commentary)

Purification (Comment) Organism
recombinant His-tagged wild-type and mutant enzymes from Escherichia coli strain BL21(DE3) to homogeneity by nickel affinity chromatgraphy Lactobacillus delbrueckii subsp. bulgaricus

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
(R)-lactate + NAD+
-
 Lactobacillus delbrueckii subsp. bulgaricus pyruvate + NADH + H+
-
 r
(R)-lactate + NAD+
-
 Lactobacillus delbrueckii subsp. bulgaricus ATCC 11842 pyruvate + NADH + H+
-
 r
2-oxobutyrate + NADH + H+
-
 Lactobacillus delbrueckii subsp. bulgaricus 2-hydroxybutyrate + NAD+
-
 ?
2-oxobutyrate + NADH + H+
-
 Lactobacillus delbrueckii subsp. bulgaricus ATCC 11842 2-hydroxybutyrate + NAD+
-
 ?
2-oxovalerate + NADH + H+
-
 Lactobacillus delbrueckii subsp. bulgaricus 2-hydroxyvalerate + NAD+
-
 ?
2-oxovalerate + NADH + H+
-
 Lactobacillus delbrueckii subsp. bulgaricus ATCC 11842 2-hydroxyvalerate + NAD+
-
 ?
additional information residues Tyr52 and Phe299 are mainly responsible for hindering larger substrates because of their short distance from the side chain of 2-oxocarboxylic acids and their steric orientation in the wild-type enzyme. Substrate specificity and enantioselectivity of D-nLDH and DnLDH mutants, overview Lactobacillus delbrueckii subsp. bulgaricus ?
-
 ?
additional information residues Tyr52 and Phe299 are mainly responsible for hindering larger substrates because of their short distance from the side chain of 2-oxocarboxylic acids and their steric orientation in the wild-type enzyme. Substrate specificity and enantioselectivity of D-nLDH and DnLDH mutants, overview Lactobacillus delbrueckii subsp. bulgaricus ATCC 11842 ?
-
 ?
phenylpyruvate + NADH + H+
-
 Lactobacillus delbrueckii subsp. bulgaricus D-phenyllactate + NAD+
-
 ?
pyruvate + NADH + H+
-
 Lactobacillus delbrueckii subsp. bulgaricus (R)-lactate + NAD+
-
 r
pyruvate + NADH + H+
-
 Lactobacillus delbrueckii subsp. bulgaricus ATCC 11842 (R)-lactate + NAD+
-
 r

Synonyms

Synonyms Comment Organism
D-nLDH
-
 Lactobacillus delbrueckii subsp. bulgaricus
NAD-dependent D-lactate dehydrogenase
-
 Lactobacillus delbrueckii subsp. bulgaricus

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
37
-
 assay at, 2-oxoacid reduction activity Lactobacillus delbrueckii subsp. bulgaricus

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
1.8
-
 phenylpyruvate pH 7.4, 37°C, recombinant His-taged mutant F299Y Lactobacillus delbrueckii subsp. bulgaricus
11.3
-
 phenylpyruvate pH 7.4, 37°C, recombinant His-taged wild-type enzyme Lactobacillus delbrueckii subsp. bulgaricus
1447
-
 phenylpyruvate pH 7.4, 37°C, recombinant His-taged mutant Y52L/F299Y Lactobacillus delbrueckii subsp. bulgaricus
2013
-
 phenylpyruvate pH 7.4, 37°C, recombinant His-taged mutant Y52L Lactobacillus delbrueckii subsp. bulgaricus

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
7.5
-
 assay at, 2-oxoacid reduction activity Lactobacillus delbrueckii subsp. bulgaricus

Cofactor

Cofactor Comment Organism Structure
NAD+
-
 Lactobacillus delbrueckii subsp. bulgaricus
NADH
-
 Lactobacillus delbrueckii subsp. bulgaricus

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
1
-
 phenylpyruvate pH 7.4, 37°C, recombinant His-taged wild-type enzyme Lactobacillus delbrueckii subsp. bulgaricus
5.7
-
 phenylpyruvate pH 7.4, 37°C, recombinant His-taged mutant F299Y Lactobacillus delbrueckii subsp. bulgaricus
1000
-
 phenylpyruvate pH 7.4, 37°C, recombinant His-taged mutant Y52L/F299Y Lactobacillus delbrueckii subsp. bulgaricus
7500
-
 phenylpyruvate pH 7.4, 37°C, recombinant His-taged mutant Y52L Lactobacillus delbrueckii subsp. bulgaricus