Glutamate dehydrogenase explained

glutamate dehydrogenase (GLDH)
Ec Number:	1.4.1.2
Cas Number:	9001-46-1
Go Code:	0004352

glutamate dehydrogenase [NAD(P)+]
Ec Number:	1.4.1.3
Cas Number:	9029-12-3
Go Code:	0004353

glutamate dehydrogenase (NADP+)
Ec Number:	1.4.1.4
Cas Number:	9029-11-2
Go Code:	0004354

K _{m} of about 1 mM), and therefore toxic levels of ammonia would have to be present in the body for the reverse reaction to proceed (that is, α-ketoglutarate and ammonia to glutamate and NAD(P)+). However, in brain, the NAD+/NADH ratio in brain mitochondria encourages oxidative deamination (i.e. glutamate to α-ketoglutarate and ammonia).^[1] In bacteria, the ammonia is assimilated to amino acids via glutamate and aminotransferases.^[2] In plants, the enzyme can work in either direction depending on environment and stress.^[3] Transgenic plants expressing microbial GLDHs are improved in tolerance to herbicide, water deficit, and pathogen infections.^[4] They are more nutritionally valuable.^[5]

The enzyme represents a key link between catabolic and anabolic pathways, and is, therefore, ubiquitous in eukaryotes. In humans the relevant genes are called GLUD1 (glutamate dehydrogenase 1) and GLUD2 (glutamate dehydrogenase 2), and there are also at least five GLDH pseudogenes in the human genome as well.^[6]

Clinical application

GLDH can be measured in a medical laboratory to evaluate the liver function. Elevated blood serum GLDH levels indicate liver damage and GLDH plays an important role in the differential diagnosis of liver disease, especially in combination with aminotransferases. GLDH is localised in mitochondria, therefore practically none is liberated in generalised inflammatory diseases of the liver such as viral hepatitides. Liver diseases in which necrosis of hepatocytes is the predominant event, such as toxic liver damage or hypoxic liver disease, are characterised by high serum GLDH levels. GLDH is important for distinguishing between acute viral hepatitis and acute toxic liver necrosis or acute hypoxic liver disease, particularly in the case of liver damage with very high aminotransferases. In clinical trials, GLDH can serve as a measurement for the safety of a drug.

Enzyme immunoassay (EIA) for glutamate dehydrogenase (GDH) can be used as screening tool for patients with Clostridioides difficile infection. The enzyme is expressed constitutively by most strains of C.diff, and can thus be easily detected in stool. Diagnosis is generally confirmed with a follow-up EIA for C. Diff toxins A and B.

Cofactors

NAD⁺ (or NADP⁺) is a cofactor for the glutamate dehydrogenase reaction, producing α-ketoglutarate and ammonium as a byproduct.^{[7] [8]}

Based on which cofactor is used, glutamate dehydrogenase enzymes are divided into the following three classes:

• EC 1.4.1.2: L-glutamate + H₂O + NAD⁺

\rightleftharpoons 2-oxoglutarate + NH₃ + NADH + H⁺

• EC 1.4.1.3: L-glutamate + H₂O + NAD(P)⁺

\rightleftharpoons 2-oxoglutarate + NH₃ + NAD(P)H + H⁺

• EC 1.4.1.4: L-glutamate + H₂O + NADP⁺

\rightleftharpoons 2-oxoglutarate + NH₃ + NADPH + H⁺

Role in flow of nitrogen

Ammonia incorporation in animals and microbes occurs through the actions of glutamate dehydrogenase and glutamine synthetase. Glutamate plays the central role in mammalian and microbe nitrogen flow, serving as both a nitrogen donor and a nitrogen acceptor.

Regulation of glutamate dehydrogenase

In humans, the activity of glutamate dehydrogenase is controlled through ADP-ribosylation, a covalent modification carried out by the gene sirt4. This regulation is relaxed in response to caloric restriction and low blood glucose. Under these circumstances, glutamate dehydrogenase activity is raised in order to increase the amount of α-ketoglutarate produced, which can be used to provide energy by being used in the citric acid cycle to ultimately produce ATP.

In microbes, the activity is controlled by the concentration of ammonium and or the like-sized rubidium ion, which binds to an allosteric site on GLDH and changes the K_m (Michaelis constant) of the enzyme.^[9]

The control of GLDH through ADP-ribosylation is particularly important in insulin-producing β cells. Beta cells secrete insulin in response to an increase in the ATP:ADP ratio, and, as amino acids are broken down by GLDH into α-ketoglutarate, this ratio rises and more insulin is secreted. SIRT4 is necessary to regulate the metabolism of amino acids as a method of controlling insulin secretion and regulating blood glucose levels.

Bovine liver glutamate dehydrogenase was found to be regulated by nucleotides in the late 1950s and early 1960s by Carl Frieden.^{[10] [11] [12] [13]} In addition to describing the effects of nucleotides like ADP, ATP and GTP he described in detail the different kinetic behavior of NADH and NADPH. As such it was one of the earliest enzymes to show what was later described as allosteric behavior.^[14]

The activation of mammalian GDH by L-leucine and some other hydrophobic amino acids has also been long known,^[15] however localization of the binding site was not clear. Only recently the new allosteric binding site for L-leucine was identified in a mammalian enzyme.^[16]

Mutations which alter the allosteric binding site of GTP cause permanent activation of glutamate dehydrogenase, and lead to hyperinsulinism-hyperammonemia syndrome.

Regulation

Allosteric regulation

This protein may use the morpheein model of allosteric regulation.^{[8] [17]}

Allosteric inhibitors:

• Guanosine triphosphate (GTP)
• Adenosine triphosphate (ATP)
• Palmitoyl-CoA
• Zn²⁺

Activators:

• Adenosine diphosphate (ADP) ^[16]
• Leucine^[16]
• l-isoleucine
• l-valine
• Guanosine diphosphate

Other Inhibitors:

• EGCG^[18]

Additionally, Mice GLDH shows substrate inhibition by which GLDH activity decreases at high glutamate concentrations.^[8]

Isozymes

Humans express the following glutamate dehydrogenase isozymes:

glutamate dehydrogenase 1 Hgncid: 4335 Symbol: GLUD1 Altsymbols: GLUD Entrezgene: 2746 Omim: 138130 Refseq: NM_005271 Uniprot: P00367 Ecnumber: 1.4.1.3 Chromosome: 10 Arm: q Band: 21.1 Locussupplementarydata: -24.3

glutamate dehydrogenase 2 Hgncid: 4336 Symbol: GLUD2 Altsymbols: GLUDP1 Entrezgene: 2747 Omim: 300144 Refseq: NM_012084 Uniprot: P49448 Ecnumber: 1.4.1.3 Chromosome: X Arm: q2 Band: 5

See also

• Anaplerotic reactions

Notes and References

1. Book: McKenna MC, Ferreira GC . Enzyme Complexes Important for the Glutamate–Glutamine Cycle . Advances in Neurobiology . The Glutamate/GABA-Glutamine Cycle . 13 . 59–98 . 2016 . 27885627 . 10.1007/978-3-319-45096-4_4 . 978-3-319-45094-0 .
2. Lightfoot DA, Baron AJ, Wootton JC . Expression of the Escherichia coli glutamate dehydrogenase gene in the cyanobacterium Synechococcus PCC6301 causes ammonium tolerance . Plant Molecular Biology . 11 . 3 . 335–44 . May 1988 . 24272346 . 10.1007/BF00027390 . 21845538 .
3. Mungur R, Glass AD, Goodenow DB, Lightfoot DA . Metabolite fingerprinting in transgenic Nicotiana tabacum altered by the Escherichia coli glutamate dehydrogenase gene . Journal of Biomedicine & Biotechnology . 2005 . 2 . 198–214 . June 2005 . 16046826 . 1184043 . 10.1155/JBB.2005.198 . free .
4. Lightfoot DA, Bernhardt K, Mungur R, Nolte S, Ameziane R, Colter A, Jones K, Iqbal MJ, Varsa E, Young B . 2007 . Improved drought tolerance of transgenic Zea mays plants that express the glutamate dehydrogenase gene (gdhA) of E. coli . Euphytica . 156 . 1–2 . 103–116 . 10.1007/s10681-007-9357-y. 11806853 .
5. Book: Wood, Andrew . Matthew A. Jenks . Lightfoot DA . Genes for use in improving nitrogen use efficiency in crops . Genes for Plant Abiotic Stress . Wiley-Blackwell . 2009 . 167–182 . 978-0-8138-1502-2 .
6. Aleshina YA, Aleshin VA. Evolutionary Changes in Primate Glutamate Dehydrogenases 1 and 2 Influence the Protein Regulation by Ligands, Targeting and Posttranslational Modifications. International Journal of Molecular Sciences. 25. 8. 2024. 10.3390/ijms25084341. Art. No. 4341. free. 38673928. 11050691.
7. Grabowska A, Nowicki M, Kwinta J . Glutamate dehydrogenase of the germinating triticale seeds: gene expression, activity distribution and kinetic characteristics . Acta Physiol. Plant. . 33 . 10.1007/s11738-011-0801-1 . 2011 . 5 . 1981–90 . free .
8. Botman D, Tigchelaar W, Van Noorden CJ . Determination of glutamate dehydrogenase activity and its kinetics in mouse tissues using metabolic mapping (quantitative enzyme histochemistry) . The Journal of Histochemistry and Cytochemistry . 62 . 11 . 802–12 . November 2014 . 25124006 . 10.1369/0022155414549071 . 4230541.
9. Wootton JC . Re-assessment of ammonium-ion affinities of NADP-specific glutamate dehydrogenases. Activation of the Neurospora crassa enzyme by ammonium and rubidium ions . The Biochemical Journal . 209 . 2 . 527–31 . February 1983 . 6221721 . 1154121 . 10.1042/bj2090527.
10. Frieden C . Glutamic dehydrogenase. II. The effect of various nucleotides on the association-dissociation and kinetic properties . The Journal of Biological Chemistry . 234 . 4 . 815–20 . April 1959 . 10.1016/S0021-9258(18)70181-6 . 13654269 . free .
11. Frieden C . The unusual inhibition of glutamate dehydrogenase by guanosine di- and triphosphate . Biochimica et Biophysica Acta . 59 . 2 . 484–6 . May 1962 . 13895207 . 10.1016/0006-3002(62)90204-4 .
12. Book: Frieden C . L-Glutamate Dehydrogenase, in The Enzymes, Vol VII . Academic Press . 3–24. 1963.
13. Frieden C . Glutamate Dehydrogenase. VI. Survey of Purine Nucleotide and Other Effects on the Enzyme from Various Sources . The Journal of Biological Chemistry . 240 . 2028–35 . May 1965 . 5 . 10.1016/S0021-9258(18)97420-X . 14299621 . free .
14. Monod J, Wyman J, Changeux JP . On the Nature of Allosteric Transitions: A Plausible Model . J Mol Biol . 12 . 88–118 . 1965 . 14343300. 10.1016/s0022-2836(65)80285-6.
15. YIELDING KL, TOMKINS GM . An effect of L-leucine and other essential amino acids on the structure and activity of glutamic dehydrogenase . Proc Natl Acad Sci U S A . 47 . 983–9 . 1961 . 7 . 13787322. 10.1073/pnas.47.7.983. 221313 . 1961PNAS...47..983L . free .
16. Aleshin VA, Bunik VI, Bruch EM, Bellinzoni M . Structural Basis for the Binding of Allosteric Activators Leucine and ADP to Mammalian Glutamate Dehydrogenase . Int J Mol Sci . 23 . 11306 . 2022 . 19 . 36232607. 10.3390/ijms231911306. 9570180 . free .
17. Selwood T, Jaffe EK . Dynamic dissociating homo-oligomers and the control of protein function . Archives of Biochemistry and Biophysics . 519 . 2 . 131–43 . March 2012 . 22182754 . 3298769 . 10.1016/j.abb.2011.11.020 .
18. Pournourmohammadi S, Grimaldi M, Stridh MH, Lavallard V, Waagepetersen HS, Wollheim CB, Maechler P . Epigallocatechin-3-gallate (EGCG) activates AMPK through the inhibition of glutamate dehydrogenase in muscle and pancreatic ß-cells: A potential beneficial effect in the pre-diabetic state? . The International Journal of Biochemistry & Cell Biology . 88 . 220–225 . July 2017 . 28137482 . 10.1016/j.biocel.2017.01.012 .