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Literature summary extracted from

  • Ochsner, A.M.; Mueller, J.E.; Mora, C.A.; Vorholt, J.A.
    In vitro activation of NAD-dependent alcohol dehydrogenases by Nudix hydrolases is more widespread than assumed (2014), FEBS Lett., 588, 2993-2999 .
    View publication on PubMed

Activating Compound

EC Number Activating Compound Comment Organism Structure
1.1.1.244 Act activator protein, UniProt ID I3EA59, the in vitro activity of Bacillus methanolicus Mdh is increased by the endogenous activator protein Act, a Nudix hydrolase, in a pH-dependent manner. Act increases the Vmax of the enzyme and decreases the Km for methanol, the increase was 23fold at pH 7.4 and 50fold at pH 9.0 Bacillus methanolicus
1.1.1.244 activator protein ACT UniProt ID I3EA59, the in vitro activity of Bacillus methanolicus Mdh is increased by the endogenous activator protein Act, a Nudix hydrolase, in a pH-dependent manner. Act increases the Vmax of the enzyme and decreases the Km for methanol, the increase is 20fold at pH 7.4 and 9fold at pH 9.0 Bacillus methanolicus
1.1.1.244 additional information the enzyme is activated by several Nudix hydrolases, relative activities of Adh enzymes, overview Bacillus methanolicus
1.1.1.244 NudBC a Nudix hydrolase from Bacillus coagulans, UniProt ID G2TKZ3 Bacillus methanolicus
1.1.1.244 NudDK a Nudix hydrolase from Desulfotomaculum kuznetsovii, UniProt ID F6CJI1 Bacillus methanolicus
1.1.1.244 NudLF a Nudix hydrolase from Lysinibacillus fusiformis, UniProt ID D7WXY3 Bacillus methanolicus
1.1.1.244 NudLS a Nudix hydrolase from Lysinibacillus sphaericus, UniProt ID B1HRL7 Bacillus methanolicus

Cloned(Commentary)

EC Number Cloned (Comment) Organism
1.1.1.244 gene mdh, sequence comparisons and phylogenetic tree, recombinant expression of His-tagged enzyme in Escherichia coli strain BL21(DE3) Bacillus methanolicus
1.1.1.244 gene mdh2, sequence comparisons and phylogenetic tree, recombinant expression of His-tagged enzyme in Escherichia coli strain BL21(DE3) Bacillus methanolicus

Protein Variants

EC Number Protein Variants Comment Organism
1.1.1.244 D38G site-directed mutagenesis, the mutant is active with NADP+ and NAD+ in contrast to the wild-type enzyme Bacillus methanolicus
1.1.1.244 D41G site-directed mutagenesis, the mutant is active with NADP+ and NAD+ in contrast to the wild-type enzyme, it shows increased activity compared to the wild-type enzyme Bacillus methanolicus
1.1.1.244 S101G site-directed mutagenesis, the mutant shows altered kinetics, reduced activity, and altered pH-dependency compared to the wild-type enzyme, overview Bacillus methanolicus
1.1.1.244 S98G site-directed mutagenesis, the mutant shows altered kinetics, reduced activity, and altered pH-dependency compared to the wild-type enzyme, overview Bacillus methanolicus

KM Value [mM]

EC Number KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
1.1.1.244 additional information
-
additional information Michaelis-Menten kinetics Bacillus methanolicus
1.1.1.244 9
-
methanol recombinant enzyme, pH 9.0, 30°C, in presence of activator Act Bacillus methanolicus
1.1.1.244 25
-
methanol recombinant enzyme, pH 7.4, 30°C, in presence of activator Act Bacillus methanolicus
1.1.1.244 51
-
ethanol recombinant enzyme, pH 7.4, 30°C, in presence of activator Act Bacillus methanolicus
1.1.1.244 96
-
methanol recombinant enzyme, pH 9.0, 30°C, in presence of activator Act Bacillus methanolicus
1.1.1.244 137
-
methanol recombinant enzyme mutant D38G, pH 7.4, 30°C, in presence of activator Act, with NADP+ Bacillus methanolicus
1.1.1.244 150
-
methanol recombinant enzyme, pH 9.0, 30°C, in absence of activator Act Bacillus methanolicus
1.1.1.244 161
-
ethanol recombinant enzyme, pH 9.0, 30°C, in presence of activator Act Bacillus methanolicus
1.1.1.244 224
-
ethanol recombinant enzyme, pH 9.0, 30°C, in absence of activator Act Bacillus methanolicus
1.1.1.244 225
-
ethanol recombinant enzyme, pH 7.4, 30°C, in absence of activator Act Bacillus methanolicus
1.1.1.244 255
-
methanol recombinant enzyme, pH 7.4, 30°C, in presence of activator Act Bacillus methanolicus
1.1.1.244 349
-
methanol recombinant enzyme, pH 7.4, 30°C, in absence of activator Act Bacillus methanolicus
1.1.1.244 379
-
methanol recombinant enzyme mutant D38G, pH 7.4, 30°C, in absence of activator Act, with NADP+ Bacillus methanolicus
1.1.1.244 416
-
methanol recombinant enzyme, pH 9.0, 30°C, in absence of activator Act Bacillus methanolicus
1.1.1.244 733
-
methanol recombinant enzyme, pH 7.4, 30°C, in absence of activator Act Bacillus methanolicus
1.1.1.244 847
-
methanol recombinant enzyme mutant S98G, pH 7.4, 30°C, in presence of activator Act Bacillus methanolicus
1.1.1.244 1151
-
methanol recombinant enzyme mutant S98G, pH 7.4, 30°C, in absence of activator Act Bacillus methanolicus

Natural Substrates/ Products (Substrates)

EC Number Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
1.1.1.244 methanol + NAD+ Bacillus methanolicus
-
formaldehyde + NADH + H+
-
r
1.1.1.244 methanol + NAD+ Bacillus methanolicus MGA3
-
formaldehyde + NADH + H+
-
r

Organism

EC Number Organism UniProt Comment Textmining
1.1.1.244 Bacillus methanolicus I3DTM5
-
-
1.1.1.244 Bacillus methanolicus I3E2P9
-
-
1.1.1.244 Bacillus methanolicus MGA3 I3DTM5
-
-
1.1.1.244 Bacillus methanolicus MGA3 I3E2P9
-
-

Purification (Commentary)

EC Number Purification (Comment) Organism
1.1.1.244 recombinant His-tagged enzyme from Escherichia coli strain BL21(DE3) by nickel affinity chromatography Bacillus methanolicus

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
1.1.1.244 ethanol + NAD+
-
Bacillus methanolicus acetaldehyde + NADH + H+
-
r
1.1.1.244 ethanol + NAD+
-
Bacillus methanolicus MGA3 acetaldehyde + NADH + H+
-
r
1.1.1.244 methanol + NAD+
-
Bacillus methanolicus formaldehyde + NADH + H+
-
r
1.1.1.244 methanol + NAD+
-
Bacillus methanolicus MGA3 formaldehyde + NADH + H+
-
r
1.1.1.244 methanol + NADP+ Mdh enzyme mutant D41G, no activity of the wild-type enzyme with NADP(H) Bacillus methanolicus formaldehyde + NADPH + H+
-
r
1.1.1.244 methanol + NADP+ Mdh enzyme mutant D98G, no activity of the wild-type enzyme with NADP(H) Bacillus methanolicus formaldehyde + NADPH + H+
-
r
1.1.1.244 methanol + NADP+ Mdh enzyme mutant D98G, no activity of the wild-type enzyme with NADP(H) Bacillus methanolicus MGA3 formaldehyde + NADPH + H+
-
r
1.1.1.244 methanol + NADP+ Mdh enzyme mutant D41G, no activity of the wild-type enzyme with NADP(H) Bacillus methanolicus MGA3 formaldehyde + NADPH + H+
-
r
1.1.1.244 additional information poor activity with 2-propanol and 1,2-popanediol Bacillus methanolicus ?
-
?
1.1.1.244 additional information poor activity with 2-propanol and 1,2-popanediol Bacillus methanolicus MGA3 ?
-
?
1.1.1.244 n-butanol + NAD+
-
Bacillus methanolicus butyraldehyde + NADH + H+
-
r
1.1.1.244 n-butanol + NAD+
-
Bacillus methanolicus MGA3 butyraldehyde + NADH + H+
-
r
1.1.1.244 n-propanol + NAD+
-
Bacillus methanolicus propionaldehyde + NADH + H+
-
r

Synonyms

EC Number Synonyms Comment Organism
1.1.1.244 MDH
-
Bacillus methanolicus
1.1.1.244 MDH2
-
Bacillus methanolicus
1.1.1.244 NAD-dependent methanol dehydrogenase
-
Bacillus methanolicus

Temperature Optimum [°C]

EC Number Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
1.1.1.244 30
-
assay at Bacillus methanolicus

pH Optimum

EC Number pH Optimum Minimum pH Optimum Maximum Comment Organism
1.1.1.244 9
-
-
Bacillus methanolicus

Cofactor

EC Number Cofactor Comment Organism Structure
1.1.1.244 NAD+
-
Bacillus methanolicus

General Information

EC Number General Information Comment Organism
1.1.1.244 evolution the enzyme belongs to the type III alcohol dehydrogenase (Adh) family Bacillus methanolicus

kcat/KM [mM/s]

EC Number kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
1.1.1.244 0.00004
-
methanol recombinant enzyme, pH 7.4, 30°C, in absence of activator Act Bacillus methanolicus
1.1.1.244 0.0001
-
methanol recombinant enzyme mutant D38G, pH 7.4, 30°C, in absence of activator Act, with NADP+ Bacillus methanolicus
1.1.1.244 0.0003
-
methanol recombinant enzyme mutant S98G, pH 7.4, 30°C, in absence of activator Act Bacillus methanolicus
1.1.1.244 0.0003
-
methanol recombinant enzyme, pH 7.4, 30°C, in absence of activator Act Bacillus methanolicus
1.1.1.244 0.0003
-
methanol recombinant enzyme, pH 9.0, 30°C, in absence of activator Act Bacillus methanolicus
1.1.1.244 0.0006
-
methanol recombinant enzyme mutant D38G, pH 7.4, 30°C, in presence of activator Act, with NADP+ Bacillus methanolicus
1.1.1.244 0.0006
-
ethanol recombinant enzyme, pH 7.4, 30°C, in absence of activator Act Bacillus methanolicus
1.1.1.244 0.0007
-
methanol recombinant enzyme mutant S98G, pH 7.4, 30°C, in presence of activator Act Bacillus methanolicus
1.1.1.244 0.0007
-
methanol recombinant enzyme, pH 9.0, 30°C, in absence of activator Act Bacillus methanolicus
1.1.1.244 0.0008
-
methanol recombinant enzyme, pH 7.4, 30°C, in presence of activator Act Bacillus methanolicus
1.1.1.244 0.0014
-
ethanol recombinant enzyme, pH 9.0, 30°C, in absence of activator Act Bacillus methanolicus
1.1.1.244 0.0028
-
methanol recombinant enzyme, pH 9.0, 30°C, in presence of activator Act Bacillus methanolicus
1.1.1.244 0.0068
-
methanol recombinant enzyme, pH 7.4, 30°C, in presence of activator Act Bacillus methanolicus
1.1.1.244 0.0074
-
ethanol recombinant enzyme, pH 7.4, 30°C, in presence of activator Act Bacillus methanolicus
1.1.1.244 0.0143
-
ethanol recombinant enzyme, pH 9.0, 30°C, in presence of activator Act Bacillus methanolicus
1.1.1.244 0.0353
-
methanol recombinant enzyme, pH 9.0, 30°C, in presence of activator Act Bacillus methanolicus