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Literature summary for 1.4.1.2 extracted from

  • Munawar, N.; Engel, P.C.
    Overexpression in a non-native halophilic host and biotechnological potential of NAD+-dependent glutamate dehydrogenase from Halobacterium salinarum strain NRC-36014 (2012), Extremophiles, 16, 463-476.
    View publication on PubMed

Activating Compound

Activating Compound Comment Organism Structure
DMSO activates Halobacterium salinarum
glycerol activates at 10% Halobacterium salinarum

Cloned(Commentary)

Cloned (Comment) Organism
overexpression in halophilic host Haloferax volcanii strain DS70 of soluble, active enzyme with NAD+-GDH activity about 800times higher in the induced sample as compared to controls, expression in inclusion bodies in mesophilic Escherichia coli Halobacterium salinarum

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.000011
-
L-glutamate pH 9.0, 40°C, recombinant enzyme Halobacterium salinarum
0.000025
-
L-glutamate pH 9.0, 40°C, recombinant enzyme in presence of DMSO Halobacterium salinarum
0.00013
-
NH3 pH 8.5, 40°C, recombinant enzyme in presence of DMSO Halobacterium salinarum
0.00035
-
NH3 pH 8.5, 40°C, recombinant enzyme Halobacterium salinarum
0.0008
-
2-oxoglutarate pH 8.5, 40°C, recombinant enzyme in presence of DMSO Halobacterium salinarum
0.0034
-
2-oxoglutarate pH 8.5, 40°C, recombinant enzyme Halobacterium salinarum
0.009
-
NADH pH 8.5, 40°C, recombinant enzyme in presence of DMSO Halobacterium salinarum
0.014
-
NADH pH 8.5, 40°C, recombinant enzyme Halobacterium salinarum
0.112
-
NAD+ pH 9.0, 40°C, recombinant enzyme in presence of DMSO Halobacterium salinarum
0.118
-
NAD+ pH 9.0, 40°C, recombinant enzyme Halobacterium salinarum

Metals/Ions

Metals/Ions Comment Organism Structure
betaine high concentrations of salt may be substituted with 30% DMSO or betaine with good stability and activity Halobacterium salinarum

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
L-glutamate + H2O + NAD+ Halobacterium salinarum
-
2-oxoglutarate + NH3 + NADH + H+
-
r
L-glutamate + H2O + NAD+ Halobacterium salinarum NRC-36014
-
2-oxoglutarate + NH3 + NADH + H+
-
r

Organic Solvent Stability

Organic Solvent Comment Organism
2-propanol GDHX retains 33% activity in 10% after 72 h, pH 8.5, room temperature. GDHX loses 94% of activity within 3 days in 10% solution Halobacterium salinarum
Acetone GDHX retains 38% activity in 10% after 72 h, pH 8.5, room temperature Halobacterium salinarum
acetonitrile GDHX retains 30% activity in 10% after 72 h, pH 8.5, room temperature Halobacterium salinarum
dimethylformamide 51% activity remaining after 72 h, pH 8.5, room temperature Halobacterium salinarum
dimethylformamide GDHX retains 73% activity in 10% DMF after 30 min at pH 8.5, 40°C, and 51% after 72 h, pH 8.5, room temperature Halobacterium salinarum
DMSO GDHX retains 100% activity in the aqueous DMSO mixtures, high concentrations of salt may be substituted with 30% DMSO or betaine with good stability and activity Halobacterium salinarum
Ethanol GDHX retains 34% activity in 10% after 72 h, pH 8.5, room temperature Halobacterium salinarum
Glycerol GDHX retains 73% activity in 10% glycerol after 72 h, pH 8.5, room temperature Halobacterium salinarum
Methanol GDHX retains more than 73% activity in 10% after 72 h, pH 8.5, room temperature Halobacterium salinarum

Organism

Organism UniProt Comment Textmining
Halobacterium salinarum P29051
-
-
Halobacterium salinarum NRC-36014 P29051
-
-

Purification (Commentary)

Purification (Comment) Organism
recombinant soluble enzyme 11.3fold from Haloferax volcanii strain DS70, by hydrophobic interaction and anion exchange chromatography, method optimization Halobacterium salinarum

Renatured (Commentary)

Renatured (Comment) Organism
enzyme solubulization from inclusion bodies after expressionin Escherichia coli in 50 mM HEPES-NaOH, pH 7.5, containing 8 M urea and 25 mM DTT, and renaturation of protein using both rapid dilution and gradual dialysis methods Halobacterium salinarum

Specific Activity [micromol/min/mg]

Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
6.52
-
recombinant enzyme in Haloferax volcanii, pH 8.5, 40°C Halobacterium salinarum
35
-
recombinant enzyme purified from Haloferax volcanii, pH 8.5, 40°C Halobacterium salinarum

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
L-glutamate + H2O + NAD+
-
Halobacterium salinarum 2-oxoglutarate + NH3 + NADH + H+
-
r
L-glutamate + H2O + NAD+
-
Halobacterium salinarum NRC-36014 2-oxoglutarate + NH3 + NADH + H+
-
r

Synonyms

Synonyms Comment Organism
NAD+-dependent GDHX
-
Halobacterium salinarum
NAD+-dependent glutamate dehydrogenase
-
Halobacterium salinarum

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
40
-
assay at Halobacterium salinarum

Temperature Range [°C]

Temperature Minimum [°C] Temperature Maximum [°C] Comment Organism
40 90 temperature profile, overview Halobacterium salinarum

Temperature Stability [°C]

Temperature Stability Minimum [°C] Temperature Stability Maximum [°C] Comment Organism
40 70 recombinant purified enzyme, fully stable at 40°C Halobacterium salinarum
80
-
recombinant purified enzyme, 30 min, 88% remaining Halobacterium salinarum
90
-
recombinant purified enzyme, 30 min, 56% remaining Halobacterium salinarum

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
15
-
NAD+ pH 9.0, 40°C, recombinant enzyme in presence of DMSO Halobacterium salinarum
16
-
NAD+ pH 9.0, 40°C, recombinant enzyme Halobacterium salinarum
17
-
L-glutamate pH 9.0, 40°C, recombinant enzyme in presence of DMSO Halobacterium salinarum
24
-
L-glutamate pH 9.0, 40°C, recombinant enzyme Halobacterium salinarum
39
-
NADH pH 8.5, 40°C, recombinant enzyme Halobacterium salinarum
46
-
2-oxoglutarate pH 8.5, 40°C, recombinant enzyme in presence of DMSO Halobacterium salinarum
47
-
NADH pH 8.5, 40°C, recombinant enzyme in presence of DMSO Halobacterium salinarum
56
-
2-oxoglutarate pH 8.5, 40°C, recombinant enzyme Halobacterium salinarum
84
-
NH3 pH 8.5, 40°C, recombinant enzyme in presence of DMSO Halobacterium salinarum
85
-
NH3 pH 8.5, 40°C, recombinant enzyme Halobacterium salinarum

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
8.5
-
amination Halobacterium salinarum
9
-
deamination Halobacterium salinarum

pH Range

pH Minimum pH Maximum Comment Organism
6.5 10.5 pH profile, overvew Halobacterium salinarum

Cofactor

Cofactor Comment Organism Structure
NAD+
-
Halobacterium salinarum
NADH
-
Halobacterium salinarum

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
0.0018
-
L-glutamate pH 9.0, 40°C, recombinant enzyme in presence of DMSO Halobacterium salinarum
0.0035
-
L-glutamate pH 9.0, 40°C, recombinant enzyme Halobacterium salinarum
0.72
-
NAD+ pH 9.0, 40°C, recombinant enzyme Halobacterium salinarum
0.75
-
NAD+ pH 9.0, 40°C, recombinant enzyme in presence of DMSO Halobacterium salinarum
1.7
-
NADH pH 8.5, 40°C, recombinant enzyme Halobacterium salinarum
1.9
-
NADH pH 8.5, 40°C, recombinant enzyme in presence of DMSO Halobacterium salinarum
16.5
-
2-oxoglutarate pH 8.5, 40°C, recombinant enzyme Halobacterium salinarum
45
-
NH3 pH 8.5, 40°C, recombinant enzyme Halobacterium salinarum
57.5
-
2-oxoglutarate pH 8.5, 40°C, recombinant enzyme in presence of DMSO Halobacterium salinarum
115
-
NH3 pH 8.5, 40°C, recombinant enzyme in presence of DMSO Halobacterium salinarum