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Literature summary extracted from

  • Tomita, T.; Kuzuyama, T.; Nishiyama, M.
    Structural basis for leucine-induced allosteric activation of glutamate dehydrogenase (2011), J. Biol. Chem., 286, 37406-37413.
    View publication on PubMedView publication on EuropePMC

Activating Compound

EC Number Activating Compound Comment Organism Structure
1.4.1.3 leucine the enzyme is subject to allosteric activation by leucine invlving ARg134, and Asp185, structural mechanism, overview Homo sapiens
1.4.1.3 leucine the enzyme is subject to allosteric activation by leucine, structural mechanism, overview Thermus thermophilus

Cloned(Commentary)

EC Number Cloned (Comment) Organism
1.4.1.3 expression of His-tagged GDH2 wild-type and mutants in Escherichia coli Homo sapiens
1.4.1.3 expression of His-tagged wild-type and mutant GDHA and GDHB in Escherichia coli strain BL21-CodonPlus (DE3)-RIL, and of untagged proteins Thermus thermophilus

Crystallization (Commentary)

EC Number Crystallization (Comment) Organism
1.4.1.3 purified recombinant untagged GDHA and GDHB in complex with each other and leucine, and GDHB in complex with glutamate, hanging drop vapor diffusion method using 0.1 M HEPES-NaOH, pH 7.0, and 0.6 M ammonium phosphate as the reservoir solution, X-ray diffraction structure determination and analysis at 2.6 A and 2.1 A resolution, respectively, molecular replacement Thermus thermophilus

Protein Variants

EC Number Protein Variants Comment Organism
1.4.1.3 A72D site-directed mutagenesis, the mutant shows no activation by leucine in contrast to the wild-type enzyme Thermus thermophilus
1.4.1.3 D166A site-directed mutagenesis, the mutant shows reduced activation by leucine compared to the wild-type enzyme Thermus thermophilus
1.4.1.3 D185A site-directed mutagenesis, the mutant shows no activation by leucine in contrast to the wild-type enzyme Homo sapiens
1.4.1.3 I71T site-directed mutagenesis, the mutant shows reduced activation by leucine compared to the wild-type enzyme Thermus thermophilus
1.4.1.3 R134A site-directed mutagenesis, the mutant shows no activation by leucine in contrast to the wild-type enzyme Thermus thermophilus
1.4.1.3 R151M site-directed mutagenesis, the mutant shows reduced activation by leucine compared to the wild-type enzyme Homo sapiens
1.4.1.3 Y38S site-directed mutagenesis, the mutant shows reduced activation by leucine compared to the wild-type enzyme Thermus thermophilus

Natural Substrates/ Products (Substrates)

EC Number Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
1.4.1.3 L-glutamate + H2O + NAD+ Thermus thermophilus
-
 2-oxoglutarate + NH3 + NADH + H+
-
 r
1.4.1.3 L-glutamate + H2O + NAD+ Homo sapiens
-
 2-oxoglutarate + NH3 + NADH + H+
-
 r
1.4.1.3 L-glutamate + H2O + NADP+ Thermus thermophilus
-
 2-oxoglutarate + NH3 + NADPH + H+
-
 r
1.4.1.3 L-glutamate + H2O + NADP+ Homo sapiens
-
 2-oxoglutarate + NH3 + NADPH + H+
-
 r

Organism

EC Number Organism UniProt Comment Textmining
1.4.1.3 Homo sapiens
-
-
-
1.4.1.3 Thermus thermophilus
-
-
-

Purification (Commentary)

EC Number Purification (Comment) Organism
1.4.1.3 recombinant His-tagged wild-type and mutant GDH2 from Escherichia coli Homo sapiens
1.4.1.3 recombinant His-tagged wild-type and mutant GDHA and GDHB from Escherichia coli strain BL21-CodonPlus (DE3)-RIL by affinity chromatography and gel filtration, and of untagged proteins by heat treatment at 90°C for 20 min, hydrophobic interaction and anion exchange chromatography, and gel filtration to homogeneity Thermus thermophilus

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
1.4.1.3 L-glutamate + H2O + NAD+
-
 Thermus thermophilus 2-oxoglutarate + NH3 + NADH + H+
-
 r
1.4.1.3 L-glutamate + H2O + NAD+
-
 Homo sapiens 2-oxoglutarate + NH3 + NADH + H+
-
 r
1.4.1.3 L-glutamate + H2O + NADP+
-
 Thermus thermophilus 2-oxoglutarate + NH3 + NADPH + H+
-
 r
1.4.1.3 L-glutamate + H2O + NADP+
-
 Homo sapiens 2-oxoglutarate + NH3 + NADPH + H+
-
 r

Subunits

EC Number Subunits Comment Organism
1.4.1.3 dimer the GdhA-GdhB-Leu complex is crystallized as a heterohexamer composed of four GdhA subunits and two GdhB subunits. In this complex, six leucine molecules are bound at subunit interfaces identified as glutamate-binding sites in the GdhB-Glu complex Thermus thermophilus
1.4.1.3 dimer the GdhA-GdhB-Leu complex is crystallized as a heterohexamer composed of four GdhA subunits and two GdhB subunits. In this complex, six leucine molecules are bound at subunit interfaces identified as glutamate-binding sites in the GdhB-Glu complex Homo sapiens
1.4.1.3 hexamer the GdhB-Glu complex is a hexamer that binds 12 glutamate molecules: six molecules are bound at the substrate-binding sites, and the remaining six are bound at subunit interfaces, each composed of three subunits Thermus thermophilus
1.4.1.3 hexamer the GdhB-Glu complex is a hexamer that binds 12 glutamate molecules: six molecules are bound at the substrate-binding sites, and the remaining six are bound at subunit interfaces, each composed of three subunits Homo sapiens
1.4.1.3 More Thermus thermophilus, possesses GDH with a unique subunit configuration composed of two different subunits, GdhA, the regulatory subunit, and GdhB, the catalytic subunit, structure of the GdhA-GdhB-Leu complex, overview Thermus thermophilus
1.4.1.3 More Thermus thermophilus, possesses GDH with a unique subunit configuration composed of two different subunits, GdhA, the regulatory subunit, and GdhB, the catalytic subunit, structure of the GdhA-GdhB-Leu complex, overview Homo sapiens

Synonyms

EC Number Synonyms Comment Organism
1.4.1.3 GDH
-
 Thermus thermophilus
1.4.1.3 GDH
-
 Homo sapiens

pH Optimum

EC Number pH Optimum Minimum pH Optimum Maximum Comment Organism
1.4.1.3 7
-
 assay at Thermus thermophilus
1.4.1.3 7
-
 assay at Homo sapiens

Cofactor

EC Number Cofactor Comment Organism Structure
1.4.1.3 NAD+
-
 Thermus thermophilus
1.4.1.3 NAD+
-
 Homo sapiens
1.4.1.3 NADH
-
 Thermus thermophilus
1.4.1.3 NADH
-
 Homo sapiens
1.4.1.3 NADP+
-
 Thermus thermophilus
1.4.1.3 NADP+
-
 Homo sapiens
1.4.1.3 NADPH
-
 Thermus thermophilus
1.4.1.3 NADPH
-
 Homo sapiens

General Information

EC Number General Information Comment Organism
1.4.1.3 additional information glutamate is bound to the active site of GdhB, the GdhB-Glu complex takes an open-like structure. No substrate is found in the active site of the GdhAGdhB-Leu complex, while six leucine molecules are found at the interfaces of three subunits Thermus thermophilus