Activating Compound | Comment | Organism | Structure |
---|---|---|---|
AMP | AMP activated GDH activity of the ternary complex, but not the GdhA-GdhB binary complex. APRTh mediates the allosteric activation of GDH by AMP. Reductive amination and oxidative deamination activity is enhanced by up to 6.3 and 2.5fold, respectively, by the addition of 1 mM AMP. | Thermus thermophilus | |
APRTh | TTC1249 (APRTh), a 23-30 kDa protein, which is annotated as adenine phosphoribosyltransferase but lacks the enzymatic activity, forms a ternary complex with the enzyme heterodimer, heterocomplex formation and structure, interaction analysis and function, detailed overview. APRTh mediates the allosteric activation of GDH by AMP | Thermus thermophilus | |
L-leucine | the crystal structure of GdhA/GdhB in a complex with leucine and biochemical analysis reveals that leucine is bound to the pockets at the GdhA-GdhA, GdhA-GdhB, and GdhB-GdhB interfaces. leucine binding increases the turnover of the GDH reaction, possibly through acceleration of the open-close cycle of the active-site cleft between the catalytic domain and NAD-binding domains | Thermus thermophilus | |
additional information | in reductive amination, the influence of AMP and leucine on the Km for NADH, 2-oxoglutarate, and ammonium is not large, although approximately 4.5, 5.3, and 10.2fold increases in Km for 2-OG are observed with AMP, leucine, and the copresence of AMP and leucine, respectively. Activation profiles of GDH activity of the TtGDH-APRTh complex by leucine and AMP, overview | Thermus thermophilus |
Cloned (Comment) | Organism |
---|---|
two GDH homologues, gdhA and gdhB, with 46% amino acid sequence identity, are present in a tandem manner as genes TTC1212 and TTC1211 on the genome of Thermus thermophilus, recombinant expression of the His-tagged ternary complex TtGDH-APRTh in Escherichia coli | Thermus thermophilus |
Protein Variants | Comment | Organism |
---|---|---|
additional information | growth analysis of the aprth knockout strain (Tt27DELTAAPRTh) and aprth-overexpressing strain (Tt27NStHisAPRTh) of Thermus thermophilus in minimal medium. The Tt27DELTAAPRTh strain exhibits delayed growth and requires approximately 36 h to reach the early stationary phase, whereas the wild-type strain reaches this phase after 21 h of cultivation. The overexpressing Tt27NStHisAPRTh strain exhibits better growth than even the wild-type strain | Thermus thermophilus |
Inhibitors | Comment | Organism | Structure |
---|---|---|---|
GTP | - |
Thermus thermophilus | |
NADH | allosteric enzyme regulation | Thermus thermophilus |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.003 | - |
NADH | recombinant enzyme complex TtGDH-APRTh, with L-leucine, pH and temperature not specified in the publication | Thermus thermophilus | |
0.0051 | - |
2-oxoglutarate | recombinant enzyme complex TtGDH-APRTh, pH and temperature not specified in the publication | Thermus thermophilus | |
0.0053 | - |
NADH | recombinant enzyme complex TtGDH-APRTh, with L-leucine and AMP, pH and temperature not specified in the publication | Thermus thermophilus | |
0.0058 | - |
NADH | recombinant enzyme complex TtGDH-APRTh, with AMP, pH and temperature not specified in the publication | Thermus thermophilus | |
0.008 | - |
NADH | recombinant enzyme complex TtGDH-APRTh, pH and temperature not specified in the publication | Thermus thermophilus | |
0.0229 | - |
2-oxoglutarate | recombinant enzyme complex TtGDH-APRTh, with AMP, pH and temperature not specified in the publication | Thermus thermophilus | |
0.024 | - |
NAD+ | recombinant enzyme complex TtGDH-APRTh, pH and temperature not specified in the publication | Thermus thermophilus | |
0.027 | - |
2-oxoglutarate | recombinant enzyme complex TtGDH-APRTh, with L-leucine, pH and temperature not specified in the publication | Thermus thermophilus | |
0.052 | - |
2-oxoglutarate | recombinant enzyme complex TtGDH-APRTh, with L-leucine and AMP, pH and temperature not specified in the publication | Thermus thermophilus | |
0.056 | - |
NAD+ | recombinant enzyme complex TtGDH-APRTh, with L-leucine, pH and temperature not specified in the publication | Thermus thermophilus | |
0.13 | - |
NAD+ | recombinant enzyme complex TtGDH-APRTh, with AMP, pH and temperature not specified in the publication | Thermus thermophilus | |
0.16 | - |
NAD+ | recombinant enzyme complex TtGDH-APRTh, with L-leucine and AMP, pH and temperature not specified in the publication | Thermus thermophilus | |
0.27 | - |
L-glutamate | recombinant enzyme complex TtGDH-APRTh, low Glu concentration, with AMP, pH and temperature not specified in the publication | Thermus thermophilus | |
0.45 | - |
L-glutamate | recombinant enzyme complex TtGDH-APRTh, with L-leucine, pH and temperature not specified in the publication | Thermus thermophilus | |
0.77 | - |
L-glutamate | recombinant enzyme complex TtGDH-APRTh, with L-leucine and AMP, pH and temperature not specified in the publication | Thermus thermophilus | |
1.2 | - |
L-glutamate | recombinant enzyme complex TtGDH-APRTh, low Glu concentration, pH and temperature not specified in the publication | Thermus thermophilus | |
19 | - |
L-glutamate | recombinant enzyme complex TtGDH-APRTh, high Glu concentration, with AMP, pH and temperature not specified in the publication | Thermus thermophilus | |
29 | - |
NH3 | recombinant enzyme complex TtGDH-APRTh, pH and temperature not specified in the publication | Thermus thermophilus | |
33 | - |
NH3 | recombinant enzyme complex TtGDH-APRTh, with L-leucine and AMP, pH and temperature not specified in the publication | Thermus thermophilus | |
43 | - |
NH3 | recombinant enzyme complex TtGDH-APRTh, with AMP, pH and temperature not specified in the publication | Thermus thermophilus | |
46 | - |
NH3 | recombinant enzyme complex TtGDH-APRTh, with L-leucine, pH and temperature not specified in the publication | Thermus thermophilus | |
52 | - |
L-glutamate | recombinant enzyme complex TtGDH-APRTh, high Glu concentration, pH and temperature not specified in the publication | Thermus thermophilus |
Molecular Weight [Da] | Molecular Weight Maximum [Da] | Comment | Organism |
---|---|---|---|
315000 | - |
recombinant His-tagged binary complex TtGDH, gel filtration | Thermus thermophilus |
331000 | - |
recombinant His-tagged ternary complex TtGDH-APRTh, gel filtration | Thermus thermophilus |
Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
---|---|---|---|---|---|---|
2-oxoglutarate + NH3 + NAD(P)H + H+ | Thermus thermophilus | - |
L-glutamate + H2O + NAD(P)+ | - |
r | |
L-glutamate + H2O + NAD(P)+ | Thermus thermophilus | - |
2-oxoglutarate + NH3 + NAD(P)H + H+ | - |
r |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Thermus thermophilus | Q93IM2 | subunit GdhA | - |
Purification (Comment) | Organism |
---|---|
recombinant His-tagged ternary complex TtGDH-APRTh from Escherichia coli by nickel affinity chromatography and gel filtration | Thermus thermophilus |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
2-oxoglutarate + NH3 + NAD(P)H + H+ | - |
Thermus thermophilus | L-glutamate + H2O + NAD(P)+ | - |
r | |
2-oxoglutarate + NH3 + NADH + H+ | - |
Thermus thermophilus | L-glutamate + H2O + NAD+ | - |
r | |
2-oxoglutarate + NH3 + NADPH + H+ | - |
Thermus thermophilus | L-glutamate + H2O + NADP+ | - |
r | |
L-glutamate + H2O + NAD(P)+ | - |
Thermus thermophilus | 2-oxoglutarate + NH3 + NAD(P)H + H+ | - |
r | |
L-glutamate + H2O + NAD+ | - |
Thermus thermophilus | 2-oxoglutarate + NH3 + NADH + H+ | - |
r | |
L-glutamate + H2O + NADP+ | - |
Thermus thermophilus | 2-oxoglutarate + NH3 + NADPH + H+ | - |
r |
Subunits | Comment | Organism |
---|---|---|
heterohexamer | 3 * 46328, GdhA, + 3 * 46112, GdhB, sequence calculation and SDS-PAGE | Thermus thermophilus |
More | GDH is composed of two heterologous subunits, GdhA and GdhB. GdhA and GdhB form a heterohexamer, in which GdhB acts as the catalytic subunit and GdhA acts as the regulatory subunit to sense leucine. In the heterocomplex, GdhB acts as the catalytic subunit, whereas GdhA lacks enzymatic activity and acts as the regulatory subunit for activation by leucine. TTC1249 (APRTh), which is annotated as adenine phosphoribosyltransferase but lacks the enzymatic activity, forms a ternary complex with the enzyme heterodimer. The ternary complex exhibits GDH activity that is activated by leucine, as observed for the GdhA-GdhB binary complex | Thermus thermophilus |
Synonyms | Comment | Organism |
---|---|---|
GDH | - |
Thermus thermophilus |
gdh-1 | - |
Thermus thermophilus |
gdhA_1 | - |
Thermus thermophilus |
NAD(P)-dependent GDH | - |
Thermus thermophilus |
TTC1211 | - |
Thermus thermophilus |
TTC1212 | - |
Thermus thermophilus |
TtGDH | - |
Thermus thermophilus |
Turnover Number Minimum [1/s] | Turnover Number Maximum [1/s] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.2 | - |
NADH | recombinant enzyme complex TtGDH-APRTh, pH and temperature not specified in the publication | Thermus thermophilus | |
0.23 | - |
2-oxoglutarate | recombinant enzyme complex TtGDH-APRTh, pH and temperature not specified in the publication | Thermus thermophilus | |
0.27 | - |
L-glutamate | recombinant enzyme complex TtGDH-APRTh, low Glu concentration, pH and temperature not specified in the publication | Thermus thermophilus | |
0.31 | - |
NH3 | recombinant enzyme complex TtGDH-APRTh, pH and temperature not specified in the publication | Thermus thermophilus | |
0.66 | - |
2-oxoglutarate | recombinant enzyme complex TtGDH-APRTh, with AMP, pH and temperature not specified in the publication | Thermus thermophilus | |
0.72 | - |
NADH | recombinant enzyme complex TtGDH-APRTh, with AMP, pH and temperature not specified in the publication | Thermus thermophilus | |
1.1 | - |
NH3 | recombinant enzyme complex TtGDH-APRTh, with AMP, pH and temperature not specified in the publication | Thermus thermophilus | |
1.2 | - |
L-glutamate | recombinant enzyme complex TtGDH-APRTh, low Glu concentration, with AMP, pH and temperature not specified in the publication | Thermus thermophilus | |
1.5 | - |
L-glutamate | recombinant enzyme complex TtGDH-APRTh, high Glu concentration, pH and temperature not specified in the publication | Thermus thermophilus | |
2.8 | - |
NAD+ | recombinant enzyme complex TtGDH-APRTh, pH and temperature not specified in the publication | Thermus thermophilus | |
3.7 | - |
NAD+ | recombinant enzyme complex TtGDH-APRTh, with L-leucine, pH and temperature not specified in the publication | Thermus thermophilus | |
4.4 | - |
L-glutamate | recombinant enzyme complex TtGDH-APRTh, high Glu concentration, with AMP, pH and temperature not specified in the publication | Thermus thermophilus | |
4.5 | - |
L-glutamate | recombinant enzyme complex TtGDH-APRTh, with L-leucine, pH and temperature not specified in the publication | Thermus thermophilus | |
9.1 | - |
NAD+ | recombinant enzyme complex TtGDH-APRTh, with AMP, pH and temperature not specified in the publication | Thermus thermophilus | |
9.1 | - |
NADH | recombinant enzyme complex TtGDH-APRTh, with L-leucine, pH and temperature not specified in the publication | Thermus thermophilus | |
12 | - |
NAD+ | recombinant enzyme complex TtGDH-APRTh, with L-leucine and AMP, pH and temperature not specified in the publication | Thermus thermophilus | |
13 | - |
L-glutamate | recombinant enzyme complex TtGDH-APRTh, with L-leucine and AMP, pH and temperature not specified in the publication | Thermus thermophilus | |
15 | - |
2-oxoglutarate | recombinant enzyme complex TtGDH-APRTh, with L-leucine, pH and temperature not specified in the publication | Thermus thermophilus | |
16 | - |
NH3 | recombinant enzyme complex TtGDH-APRTh, with L-leucine, pH and temperature not specified in the publication | Thermus thermophilus | |
29 | - |
NADH | recombinant enzyme complex TtGDH-APRTh, with L-leucine and AMP, pH and temperature not specified in the publication | Thermus thermophilus | |
37 | - |
2-oxoglutarate | recombinant enzyme complex TtGDH-APRTh, with L-leucine and AMP, pH and temperature not specified in the publication | Thermus thermophilus | |
44 | - |
NH3 | recombinant enzyme complex TtGDH-APRTh, with L-leucine and AMP, pH and temperature not specified in the publication | Thermus thermophilus |
Cofactor | Comment | Organism | Structure |
---|---|---|---|
NAD+ | - |
Thermus thermophilus | |
NADH | - |
Thermus thermophilus | |
NADP+ | - |
Thermus thermophilus | |
NADPH | - |
Thermus thermophilus |
General Information | Comment | Organism |
---|---|---|
evolution | GDH is a widely distributed enzyme among all domains of life. Mammalian GDH is regulated allosterically by multiple metabolites, in which the antenna helix plays a key role to transmit the allosteric signals. In contrast, bacterial GDH is believed not to be regulated allosterically because it lacks the antenna helix | Thermus thermophilus |
malfunction | growth analysis of the aprth knockout strain (Tt27DELTAAPRTh) and aprth-overexpressing strain (Tt27NStHisAPRTh) of Thermus thermophilus in minimal medium. The Tt27DELTAAPRTh strain exhibits delayed growth and requires approximately 36 h to reach the early stationary phase, whereas the wild-type strain reaches this phase after 21 h of cultivation. The overexpressing Tt27NStHisAPRTh strain exhibits better growth than even the wild-type strain | Thermus thermophilus |
additional information | glutamate dehydrogenase (GDH) from Thermus thermophilus is composed of two heterologous subunits, GdhA and GdhB. In the heterocomplex, GdhB acts as the catalytic subunit, whereas GdhA lacks enzymatic activity and acts as the regulatory subunit for activation by leucine. TTC1249 (APRTh), which is annotated as adenine phosphoribosyltransferase but lacks the enzymatic activity, forms a ternary complex with the enzyme heterodimer. The ternary complex exhibits GDH activity that is activated by leucine, as observed for the GdhA-GdhB binary complex | Thermus thermophilus |
physiological function | GDH catalyzes the synthesis and degradation of glutamate using NAD(P)(H). Bacterial GDH is believed not to be regulated allosterically because it lacks the antenna helix. TtGDH is activated by AMP in a complex with APRTh where APRTh is necessary for the AMP-mediated allosteric activation of TtGDH. L-Leucine is also required for allosteric regulation/activation of the enzyme | Thermus thermophilus |