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Literature summary for 1.1.1.76 extracted from

  • Pu, Z.; Ji, F.; Wang, J.; Zhang, Y.; Sun, W.; Bao, Y.
    Rational design of meso-2,3-butanediol dehydrogenase by molecular dynamics simulation and experimental evaluations (2017), FEBS Lett., 591, 3402-3413 .
    View publication on PubMed
Search for more literature for 1.1.1.76

Cloned(Commentary)

Cloned (Comment) Organism
gene budC, recombinant expression of SUMO-His-tagged wild-type and mutant enzymes in Escherichia coli strain BL21-Gold from plasmid pET28a-SUMO-KpBDH Klebsiella pneumoniae

Protein Variants

Protein Variants Comment Organism
F212S site-directed mutagenesis, the mutant shows highly reduced activity compared to wild-type Klebsiella pneumoniae
F212W site-directed mutagenesis, the mutant shows highly reduced activity compared to wild-type Klebsiella pneumoniae
F212Y site-directed mutagenesis, the kcat of the mutant is enhanced 4-8fold compared to wild-type Klebsiella pneumoniae
N146A site-directed mutagenesis, inactive mutant Klebsiella pneumoniae
N146Q site-directed mutagenesis, the mutant shows unaltered activity compared to wild-type Klebsiella pneumoniae

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
6.26
-
 (2S,3S)-butane-2,3-diol pH 8.0, 25°C, recombinant wild-type enzyme Klebsiella pneumoniae
7.45
-
 (2S,3S)-butane-2,3-diol pH 8.0, 25°C, recombinant mutant F212Y Klebsiella pneumoniae
10.2
-
 (2S,3S)-butane-2,3-diol pH 8.0, 25°C, recombinant mutant F212W Klebsiella pneumoniae
10.86
-
 (2S,3S)-butane-2,3-diol pH 8.0, 25°C, recombinant mutant N146Q Klebsiella pneumoniae
23.95
-
 (2S,3S)-butane-2,3-diol pH 8.0, 25°C, recombinant mutant F212S Klebsiella pneumoniae

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
(2R,3S)-butane-2,3-diol + NAD+ Klebsiella pneumoniae
-
 (3R)-acetoin + NADH + H+
-
 r
(2S,3S)-butane-2,3-diol + NAD+ Klebsiella pneumoniae
-
 (S)-acetoin + NADH + H+
-
 r

Organism

Organism UniProt Comment Textmining
Klebsiella pneumoniae Q48436
-
-

Purification (Commentary)

Purification (Comment) Organism
recombinant SUMO-His-tagged wild-type and mutant enzymes from Escherichia coli strain BL21-Gold by affinity chromatography, desalting gel filtration, and protease-mediated tag cleavage Klebsiella pneumoniae

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
(2R,3S)-butane-2,3-diol + NAD+
-
 Klebsiella pneumoniae (3R)-acetoin + NADH + H+
-
 r
(2R,3S)-butane-2,3-diol + NAD+ preferred substrate Klebsiella pneumoniae (3R)-acetoin + NADH + H+
-
 r
(2S,3S)-butane-2,3-diol + NAD+
-
 Klebsiella pneumoniae (S)-acetoin + NADH + H+
-
 r
additional information the meso-2,3-butanediol dehydrogenase from Klebsiella pneumoniae is active with meso-2,3-butanediol, but also with (2S,3S)-butane-2,3-diol converting them to (3R)-acetoin and (3S)-acetoin, respectively. Additionally the enzyme also has diacetyl reductase [(S)-acetoin forming] activity (EC 1.1.1.304) Klebsiella pneumoniae ?
-
-

Synonyms

Synonyms Comment Organism
budC
-
 Klebsiella pneumoniae

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
25
-
 assay at Klebsiella pneumoniae

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
0.56
-
 (2S,3S)-butane-2,3-diol pH 8.0, 25°C, recombinant mutant F212S Klebsiella pneumoniae
0.56
-
 (2S,3S)-butane-2,3-diol pH 8.0, 25°C, recombinant mutant F212W Klebsiella pneumoniae
0.81
-
 (2S,3S)-butane-2,3-diol pH 8.0, 25°C, recombinant mutant N146Q Klebsiella pneumoniae
3.51
-
 (2S,3S)-butane-2,3-diol pH 8.0, 25°C, recombinant wild-type enzyme Klebsiella pneumoniae
11.06
-
 (2S,3S)-butane-2,3-diol pH 8.0, 25°C, recombinant mutant F212Y Klebsiella pneumoniae

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
8
-
 assay at Klebsiella pneumoniae

Cofactor

Cofactor Comment Organism Structure
NAD+
-
 Klebsiella pneumoniae
NADH
-
 Klebsiella pneumoniae

General Information

General Information Comment Organism
evolution the enzyme belongs to the short-chain dehydrogenases/reductases Klebsiella pneumoniae
additional information identification of the the active tunnel of meso-2,3-BDH. The two short alpha-helices positioned away from the alpha4-helix possibly expose the hydrophobic ligand-binding cavity, gating the exit of product and cofactor from the activity pocket. AC binds in the active pocket including Ser139, Gln140, Ala141, Leu149, Tyr152, Gly183, Ile184, and Trp190. Residues Phe212 and Asn146 function as the key product-release sites. Three catalytic residues are Ser139, Tyr152, and Lys156. Docking study using the structure of meso-2,3-BDH (PDB ID 1GEG), molecular dynamics simulation Klebsiella pneumoniae
physiological function the meso-2,3-butanediol dehydrogenase (meso-2,3-BDH) catalyzes NAD+-dependent conversion of meso-2,3-butanediol to acetoin (AC), a crucial external energy storage molecule in fermentive bacteria. The interconversion between (3R)-AC and meso-2,3-BD or (3S)-AC and (2S,3S)-2,3-BD is catalyzed by meso-2,3-butanediol dehydrogenase (meso-2,3-BDH) Klebsiella pneumoniae

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
0.023
-
 (2S,3S)-butane-2,3-diol pH 8.0, 25°C, recombinant mutant F212S Klebsiella pneumoniae
0.055
-
 (2S,3S)-butane-2,3-diol pH 8.0, 25°C, recombinant mutant F212W Klebsiella pneumoniae
0.075
-
 (2S,3S)-butane-2,3-diol pH 8.0, 25°C, recombinant mutant N146Q Klebsiella pneumoniae
0.56
-
 (2S,3S)-butane-2,3-diol pH 8.0, 25°C, recombinant wild-type enzyme Klebsiella pneumoniae
1.49
-
 (2S,3S)-butane-2,3-diol pH 8.0, 25°C, recombinant mutant F212Y Klebsiella pneumoniae