Threonine ammonia-lyase

Last updated
L-threonine ammonia-lyase
TD Image 1.png
A 3d cartoon depiction of the threonine deaminase tetramer
Identifiers
EC number 4.3.1.19
Databases
IntEnz IntEnz view
BRENDA BRENDA entry
ExPASy NiceZyme view
KEGG KEGG entry
MetaCyc metabolic pathway
PRIAM profile
PDB structures RCSB PDB PDBe PDBsum
Gene Ontology AmiGO / QuickGO

Threonine ammonia-lyase, also commonly referred to as threonine deaminase or threonine dehydratase, is an enzyme responsible for catalyzing the conversion of L-threonine into alpha-ketobutyrate and ammonia. Alpha-ketobutyrate can be converted into L-isoleucine, so threonine ammonia-lyase functions as a key enzyme in BCAA synthesis. [1] It employs a pyridoxal-5'-phosphate cofactor, similar to many enzymes involved in amino acid metabolism. It is found in bacteria, yeast, and plants, though most research to date has focused on forms of the enzyme in bacteria. This enzyme was one of the first in which negative feedback inhibition by the end product of a metabolic pathway was directly observed and studied. [2] The enzyme serves as an excellent example of the regulatory strategies used in amino acid homeostasis.

Contents

Structure

Threonine ammonia-lyase is a tetramer of identical subunits, and is arranged as a dimer of dimers. Each subunit has two domains: a domain containing the catalytic active site and a domain with allosteric regulatory sites. The two have been shown to be distinct regions, [3] but the regulatory site of one subunit actually interacts with the catalytic site of another subunit. [4] Both domains contain the repeating structural motif of beta sheets surrounded by alpha helices. [5] While the threonine binding site is not perfectly understood, structural studies do reveal how the pyridoxal phosphate cofactor is bound. [4] The PLP cofactor is bonded to a lysine residue by means of a Schiff base, and the phosphate group of PLP is held in place by amine groups derived from a repeating sequence of glycine residues. The aromatic ring is bound to phenylalanine, and the nitrogen on the ring is hydrogen bonded to hydroxyl group-containing residues. [6]

Key residues that interact with PLP within the active site. Generated from 1VE5. TD PLP Site 2.png
Key residues that interact with PLP within the active site. Generated from 1VE5.

Mechanism

The mechanism of threonine ammonia-lyase is analogous to other deaminating PLP enzymes in its use of Schiff base intermediates. [7] Initially, the amine group of threonine attacks the lysine/PLP Schiff base, displacing lysine. After deprotonation of the amino acid alpha carbon and subsequent dehydration (hence the common name threonine dehydratase), a new Schiff base is formed. This Schiff base is replaced by lysine attack, reforming the catalytically active PLP and releasing an initial alkene-containing product. This product tautomerizes, and after hydrolysis of the Schiff base, the final products are generated. [8] [9] After the final alpha-ketobutyrate product is generated, isoleucine is synthesized by progressing through the intermediates alpha-acetohydroxybutyrate to alpha-beta-dihydroxy-beta-methylvalerate, then to alpha-keto-beta-methylvalerate. [10]

The mechanism of threonine ammonia-lyase. PLP and lysine are shown in blue. Mechanism of TD.png
The mechanism of threonine ammonia-lyase. PLP and lysine are shown in blue.

Regulation

Threonine ammonia-lyase has been shown to not follow Michaelis-Menten kinetics, rather, it is subject to complex allosteric control. [11] The enzyme is inhibited by isoleucine, the product of the pathway it participates in, and is activated by valine, the product of a parallel pathway. [1] Thus, an increase in isoleucine concentration shuts off its production, and an increase in valine concentration diverts starting material (Hydroxyethyl-TPP) away from valine production. The enzyme has two binding sites for isoleucine; one has a high affinity for isoleucine and the other has a low affinity. [12] The binding of isoleucine to the high affinity site increases the binding affinity of the low affinity site, and enzyme deactivation occurs when isoleucine binds to the low affinity site. Valine promotes enzyme activity by competitively binding to the high affinity site, preventing isoleucine from having an inhibitory effect. [12] The combination of these two feedback methods balances the concentration of BCAAs.

A diagram of the feedback regulatory pathways of threonine ammonia-lyase. Regulation of TD.png
A diagram of the feedback regulatory pathways of threonine ammonia-lyase.

Isoforms and other functions

Multiple forms of threonine ammonia-lyase have been observed in a variety of species of organism. In Escherichia coli , a system in which the enzyme has been studied extensively, two different forms of the enzyme are found. One is biosynthetic and resembles the enzyme characteristics presented here, while the other is degradative and functions to generate carbon fragments for energy production. [2] The pair of isoforms has also been observed in other bacteria. In many bacteria, the biodegradative isoform of the enzyme is expressed in anaerobic conditions and is promoted by cAMP and threonine, while the biosynthetic isoform is expressed in aerobic conditions. [13] This allows the bacterium to balance energy stores and inhibit energy-consuming synthetic pathways when energy is not abundant.

In plants, threonine ammonia-lyase is important in defense mechanisms against herbivores and is upregulated in response to abiotic stress. [14] An adapted isoform of the enzyme with unique properties that deter herbivores is expressed in plant leaves. The catalytic domain of this isoform is extremely resistant to proteolysis, while the regulatory domain degrades readily, so upon ingestion by another organism, the threonine deamination capabilities of the enzyme go unchecked. This degrades threonine before the herbivore can absorb it, starving the herbivore of an essential amino acid. [15] Studies of threonine ammonia-lyase in plants have also offered new strategies in the development of GMOs with increased nutritional value by increasing essential amino acid content. [14]

Other more exotic forms of the enzyme have been found that are extremely small in size, but still retain all catalytic and regulatory functions. [4]

Evolution

There are five major fold types for PLP-dependent enzymes. Threonine ammonia-lyase is a member of the Fold Type II family, also known as the tryptophan synthase family. [7] Though threonine ammonia-lyase does not possess substrate tunneling like tryptophan synthase does, it contains much conserved homology. Threonine ammonia-lyase is most closely related to serine dehydratase, and both possess the same general catalytic mechanism. [9] In fact, threonine ammonia-lyase has been shown to exhibit some specificity towards serine and can convert serine into pyruvate. [2] The regulatory domain of threonine ammonia-lyase is very similar to the regulatory domain of phosphoglycerate dehydrogenase. [4] All of these relationships demonstrate that threonine ammonia-lyase has close evolutionary ties to these enzymes. Due to the degree of conserved structure and sequence in enzymes that recognize amino acids, it is likely that the evolutionary diversity of these enzymes came about by the matching together of individual regulatory and catalytic domains in various ways. [1]

Relevance to humans

Threonine ammonia-lyase is not found in humans. Thus, this is one example of why humans cannot synthesize all 20 proteinogenic amino acids; in this specific case, humans cannot convert threonine into isoleucine and must consume isoleucine in the diet. [1] The enzyme has also been studied in the past as a possible tumor suppressing agent for the previously described reasons, in that it deprives tumor cells of an essential amino acid and kills them, [16] but this treatment has not been utilized.

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Cystathionine beta synthase mammalian protein found in Homo sapiens

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Serine dehydratase

Serine dehydratase or L-serine ammonia lyase (SDH) is in the β-family of pyridoxal phosphate-dependent (PLP) enzymes. SDH is found widely in nature, but its structural and chemical properties vary greatly among species. SDH is found in yeast, bacteria, and the cytoplasm of mammalian hepatocytes. The reaction it catalyzes is the deamination of L-serine to yield pyruvate, with the release of ammonia.

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Methionine gamma-lyase class of enzymes

Methionine gamma-lyase (MGL) is an enzyme in the γ-family of PLP-dependent enzymes. It degrades sulfur-containing amino acids to α-keto acids, ammonia, and thiols. Because sulfur-containing amino acids play a role in multiple biological processes, the regulation of these amino acids is essential. Additionally, it is crucial to maintain low homocysteine levels for the proper functioning of various pathways and for preventing the toxic effects of the cysteine homologue. Methionine gamma-lyase has been found in several bacteria (Clostridiums porogenes, Pseudomonas ovalis, Pseudomonas putida, Aeromonas sp., Citrobacter intermedius, Brevibacterium linens, Citrobacter freundii, Porphyromonas gingivalis, Treponema denticola), parasitic protozoa (Trichomonas vaginalis, Entamoeba histolytica), and plants (Arabidopsis thaliana).

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Arginine decarboxylase class of enzymes

Acid-Induced Arginine Decarboxylase (AdiA), also commonly referred to as arginine decarboxylase, is an enzyme responsible for catalyzing the conversion of L-arginine into agmatine and carbon dioxide. The process consumes a proton in the decarboxylation and employs a pyridoxal-5'-phosphate (PLP) cofactor, similar to other enzymes involved in amino acid metabolism, such as ornithine decarboxylase and glutamine decarboxylase. It is found in bacteria and virus, though most research has so far focused on forms of the enzyme in bacteria. During the AdiA catalyzed decarboxylation of arginine, the necessary proton is consumed from the cell cytoplasm which helps to prevent the over-accumulation of protons inside the cell and serves to increase the intracellular pH. Arginine decarboxylase is part of an enzymatic system in Escherichia coli, Salmonella Typhimurium, and methane-producing bacteria Methanococcus jannaschii that makes these organisms acid resistant and allows them to survive under highly acidic medium.

Diaminopimelate decarboxylase class of enzymes

In enzymology, diaminopimelate decarboxylase, also known as diaminopimelic acid decarboxylase, DAPDC, meso-diaminopimelate decarboxylase, DAP-decarboxylase, and meso-2,6-diaminoheptanedioate carboxy-lyase, is an enzyme that catalyzes the cleavage of carbon-carbon bonds in meso 2,6 diaminoheptanedioate to produce CO2 and L-lysine, the essential amino acid. It employs the cofactor pyridoxal phosphate, also known as PLP, which participates in numerous enzymatic transamination, decarboxylation and deamination reactions.

Cys/Met metabolism PLP-dependent enzyme family

In molecular biology, the Cys/Met metabolism PLP-dependent enzyme family is a family of proteins including enzymes involved in cysteine and methionine metabolism which use PLP (pyridoxal-5'-phosphate) as a cofactor.

References

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