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Literature summary for 1.1.1.6 extracted from

  • Beauchamp, J.; Gross, P.G.; Vieille, C.
    Characterization of Thermotoga maritima glycerol dehydrogenase for the enzymatic production of dihydroxyacetone (2014), Appl. Microbiol. Biotechnol., 98, 7039-7050.
    View publication on PubMed

Application

Application Comment Organism
biotechnology GlyDH is active with immobilized N6-CM-NAD+, suggesting that N6-CM-NAD+ can be immobilized on an electrode to allow TmGlyDH activity in a system that reoxidizes the cofactor electrocatalytically, development of a bioelectrocatalytic reactor Thermotoga maritima
synthesis GlyDH is active with immobilized N6-CM-NAD+, suggesting that N6-CM-NAD+ can be immobilized on an electrode to allow TmGlyDH activity in a system that reoxidizes the cofactor electrocatalytically, development of a bioelectrocatalytic reactor Thermotoga maritima

Cloned(Commentary)

Cloned (Comment) Organism
gene gldA, recombinant expression of C-terminally His6-tagged enzyme in Escherichia coli strain BL21(DE3) Thermotoga maritima

Protein Variants

Protein Variants Comment Organism
I154A site-directed mutagenesis, the mutant shows altered activity with cofactor derivative N6-CM-NAD+ immobilized on Sepharose beads compared to the wild-type enzyme Thermotoga maritima
I154A/K157G site-directed mutagenesis, the mutant shows altered activity with cofactor derivative N6-CM-NAD+ immobilized on Sepharose beads compared to the wild-type enzyme Thermotoga maritima
I154A/K157N site-directed mutagenesis, the mutant shows altered activity with cofactor derivative N6-CM-NAD+ immobilized on Sepharose beads compared to the wild-type enzyme Thermotoga maritima
K157G site-directed mutagenesis, the mutant shows altered activity with cofactor derivative N6-CM-NAD+ immobilized on Sepharose beads compared to the wild-type enzyme Thermotoga maritima
K157N site-directed mutagenesis, the mutant shows altered activity with cofactor derivative N6-CM-NAD+ immobilized on Sepharose beads compared to the wild-type enzyme Thermotoga maritima
V44A site-directed mutagenesis, the mutant shows altered activity with cofactor derivative N6-CM-NAD+ immobilized on Sepharose beads compared to the wild-type enzyme Thermotoga maritima
V44A/I154A site-directed mutagenesis, the mutant shows altered activity with cofactor derivative N6-CM-NAD+ immobilized on Sepharose beads compared to the wild-type enzyme Thermotoga maritima
V44A/K157G site-directed mutagenesis, the mutant shows altered activity with cofactor derivative N6-CM-NAD+ immobilized on Sepharose beads compared to the wild-type enzyme Thermotoga maritima
V44A/K157G site-directed mutagenesis, wild-type enzyme TmGlyDH shows little activity with N6-carboxymethyl-NAD+ (N6-CM-NAD), an NAD+ analogue modified for easy immobilization to amino groups, but the double mutation V44A/K157G increases catalytic efficiency with N6-CMNAD+ by 10fold Thermotoga maritima
V44A/K157N site-directed mutagenesis, the mutant shows altered activity with cofactor derivative N6-CM-NAD+ immobilized on Sepharose beads compared to the wild-type enzyme Thermotoga maritima

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
additional information
-
additional information Michaelis-Menten kinetics, cooperative behavior of TmGlyDH Thermotoga maritima
0.02
-
NADH pH 7.9, 50°C, recombinant wild-type enzyme Thermotoga maritima
0.021
-
NAD+ pH 7.4, 50°C, recombinant mutant V44A/K157G Thermotoga maritima
0.024
-
NAD+ pH 7.4, 50°C, recombinant wild-type enzyme Thermotoga maritima
0.025
-
NAD+ pH 7.4, 50°C, recombinant mutant K157N Thermotoga maritima
0.031
-
NAD+ pH 7.4, 50°C, recombinant mutant K157G Thermotoga maritima
0.039
-
NAD+ pH 7.4, 50°C, recombinant mutant V44A/K157N Thermotoga maritima
0.051
-
NAD+ pH 7.9, 50°C, recombinant wild-type enzyme Thermotoga maritima
0.07
-
NAD+ pH 7.4, 50°C, recombinant mutant V44A Thermotoga maritima
0.08
-
N6-carboxymethyl-NAD+ pH 7.4, 50°C, recombinant mutant K157G Thermotoga maritima
0.08
-
N6-carboxymethyl-NAD+ mutant enzyme K157G, at pH 7.4 and 50°C Thermotoga maritima
0.088
-
NAD+ pH 7.4, 50°C, recombinant mutant I154A Thermotoga maritima
0.108
-
NAD+ pH 7.4, 50°C, recombinant mutant I154A/K157G Thermotoga maritima
0.15
-
N6-carboxymethyl-NAD+ pH 7.4, 50°C, recombinant mutant V44A Thermotoga maritima
0.15
-
N6-carboxymethyl-NAD+ mutant enzyme V44A, at pH 7.4 and 50°C Thermotoga maritima
0.186
-
N6-carboxymethyl-NAD+ pH 7.4, 50°C, recombinant mutant V44A/K157N Thermotoga maritima
0.186
-
N6-carboxymethyl-NAD+ mutant enzyme V44A/ K157N, at pH 7.4 and 50°C Thermotoga maritima
0.25
-
N6-carboxymethyl-NAD+ pH 7.4, 50°C, recombinant mutant V44A/K157G Thermotoga maritima
0.25
-
N6-carboxymethyl-NAD+ mutant enzyme V44A/K157G, at pH 7.4 and 50°C Thermotoga maritima
0.351
-
NAD+ pH 7.4, 50°C, recombinant mutant V44A/I154A Thermotoga maritima
0.36
-
N6-carboxymethyl-NAD+ pH 7.4, 50°C, recombinant mutant K157N Thermotoga maritima
0.36
-
N6-carboxymethyl-NAD+ mutant enzyme K157N, at pH 7.4 and 50°C Thermotoga maritima
0.37
-
N6-carboxymethyl-NAD+ pH 7.4, 50°C, recombinant wild-type enzyme Thermotoga maritima
0.37
-
N6-carboxymethyl-NAD+ wild type enzyme, at pH 7.4 and 50°C Thermotoga maritima
0.6
-
NAD+ pH 7.4, 50°C, recombinant mutant V44A/I154A/K157G Thermotoga maritima
0.73
-
N6-carboxymethyl-NAD+ pH 7.4, 50°C, recombinant mutant I154A Thermotoga maritima
0.73
-
N6-carboxymethyl-NAD+ mutant enzyme I154A, at pH 7.4 and 50°C Thermotoga maritima
1.5
-
N6-carboxymethyl-NAD+ pH 7.4, 50°C, recombinant mutant I154A/K157G Thermotoga maritima
1.5
-
N6-carboxymethyl-NAD+ mutant enzyme I154A/K157G, at pH 7.4 and 50°C Thermotoga maritima
2.3
-
N6-carboxymethyl-NAD+ pH 7.4, 50°C, recombinant mutant V44A/I154A/K157G Thermotoga maritima
2.3
-
N6-carboxymethyl-NAD+ mutant enzyme V44A/I154A/K157G, at pH 7.4 and 50°C Thermotoga maritima
2.9
-
N6-carboxymethyl-NAD+ pH 7.4, 50°C, recombinant mutant V44A/I154A Thermotoga maritima
2.9
-
N6-carboxymethyl-NAD+ mutant enzyme V44A/ I154A, at pH 7.4 and 50°C Thermotoga maritima

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
glycerol + NAD+ Thermotoga maritima
-
glycerone + NADH + H+
-
r
glycerol + NAD+ Thermotoga maritima MSB8 / DSM 3109 / ATCC 43589
-
glycerone + NADH + H+
-
r

Organism

Organism UniProt Comment Textmining
Thermotoga maritima Q9WYQ4
-
-
Thermotoga maritima MSB8 / DSM 3109 / ATCC 43589 Q9WYQ4
-
-

Purification (Commentary)

Purification (Comment) Organism
gene gldA, recombinant C-terminally His6-tagged enzyme from Escherichia coli strain BL21(DE3) by nickel affinity chromatography, ultrafiltration, and dialysis Thermotoga maritima

Specific Activity [micromol/min/mg]

Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
45
-
purified recombinant enzyme, oxidation of glycerol, 80°C, pH 7.4 Thermotoga maritima

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
(R)-1,2-propanediol + NAD+ the enzyme prefers the R-enantiomer Thermotoga maritima hydroxyacetone + NADH
-
r
(R)-1,2-propanediol + NAD+ the enzyme prefers the R-enantiomer Thermotoga maritima MSB8 / DSM 3109 / ATCC 43589 hydroxyacetone + NADH
-
r
1,2-butanediol + NAD+
-
Thermotoga maritima ? + NADH
-
?
1,2-butanediol + NAD+
-
Thermotoga maritima MSB8 / DSM 3109 / ATCC 43589 ? + NADH
-
?
glycerol + N6-carboxymethyl-NAD+
-
Thermotoga maritima glycerone + N6-carboxymethyl-NADH + H+
-
r
glycerol + N6-carboxymethyl-NAD+
-
Thermotoga maritima MSB8 / DSM 3109 / ATCC 43589 glycerone + N6-carboxymethyl-NADH + H+
-
r
glycerol + N6-CM-NAD+
-
Thermotoga maritima glycerone + N6-carboxymethyl-NADH + H+
-
r
glycerol + N6-CM-NAD+
-
Thermotoga maritima MSB8 / DSM 3109 / ATCC 43589 glycerone + N6-carboxymethyl-NADH + H+
-
r
glycerol + NAD+
-
Thermotoga maritima glycerone + NADH + H+
-
r
glycerol + NAD+
-
Thermotoga maritima MSB8 / DSM 3109 / ATCC 43589 glycerone + NADH + H+
-
r

Synonyms

Synonyms Comment Organism
GldA
-
Thermotoga maritima
TmGlyDH
-
Thermotoga maritima

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
80
-
assay at Thermotoga maritima

Temperature Range [°C]

Temperature Minimum [°C] Temperature Maximum [°C] Comment Organism
90
-
reduction activity Thermotoga maritima

Temperature Stability [°C]

Temperature Stability Minimum [°C] Temperature Stability Maximum [°C] Comment Organism
50
-
the recombinant enzyme is initially more stable in the presence of DHA than in its absence, maintaining 100% activity over 120 min, but it quickly inactivates after 120 min, losing over 84% of its activity in the next 5 h Thermotoga maritima
80
-
the recombinant enzyme, 40% activity remaining after 25 min, 20% activity remaining after 150 min Thermotoga maritima

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
7.9
-
-
Thermotoga maritima

pH Range

pH Minimum pH Maximum Comment Organism
6 8.5 oxidation activity range Thermotoga maritima
6.4 9 reduction activity range Thermotoga maritima

Cofactor

Cofactor Comment Organism Structure
N6-carboxymethyl-NAD+ wild-type enzyme TmGlyDH shows little activity with N6-carboxymethyl-NAD+ (N6-CM-NAD), an NAD+ analogue modified for easy immobilization to amino groups, but the double mutation V44A/K157G increases catalytic efficiency with N6-CMNAD+ by 10fold. kinetics with N6-CM-NAD+ and modelling, overview. N6-CM-NAD+ is immobilized on cyanogen bromide-activated Sepharose 4b Thermotoga maritima
NAD+
-
Thermotoga maritima
NADH
-
Thermotoga maritima