Threonine Amino Acids: Properties, Function, Benefits, and Sources

What is threonine?

Threonine (Thr), also known as α-amino-β-hydroxybutyric acid, is a glucogenic and ketogenic necessary amino acid. Human plasma levels of threonine can be significantly raised by supplementation. Most mammals need threonine as a precursor for glycine synthesis, and studies have demonstrated that supplementing with threonine raises the levels of glycine neurotransmitters in rats' brains and plasma glycine in pigs. Pigs' rates of protein synthesis were lowered by either an excess or a shortage of dietary threonine. Age-related decreases in threonine levels were seen in the plasma of old rats. High levels of threonine (together with two other amino acids) were associated with longevity, as measured by living to be at least 80 years old, according to a metabolomics research of 647 human individuals. Threonine levels in human and mouse embryonic stem cells can control the proliferation and G1/S phase transition of the cell cycle. In mammals, a lack of threonine has been linked to neurological malfunction and depression. Supplementing hens with threonine boosted their immune response and protected them from intestinal barrier disruption and inflammation brought on by LPS. Pigs that were supplemented with threonine showed improved resistance to a viral challenge. It has been demonstrated that threonine supplementation increases yeast's chronological lifespan. Nevertheless, a different group under different conditions discovered that by blocking TOR kinase, threonine (and valine) limitation increased the chronological longevity of yeast.

Metabolism of amino acids influence one-carbon and NADPH redox metabolism. (Canfield C A., et al., 2019)

Threonine structure

The structural examination of Threonine revealed the presence of a second asymmetric carbon, in addition to the chiral a-carbon atom, which is attached to a methyl group, a hydroxy group, the a-C, and a hydrogen atom. In addition to isoleucine, this molecule is the second proteinogenic amino acid possessing two stereogenic centers. Subsequently, all combinations of the chiral carbon atoms were identified, isolated, and structurally described. It possesses a second asymmetrical carbon atom on its side chain, resulting in four isomers, of which only L-threonine is found in proteins. The hydroxyl group, exemplified by serine, engages in phosphorylation and dephosphorylation events as well as interactions with sugar residues. Phosphorylation can generate phosphoserine and phosphothreonine from the hydroxyl group of particular serine and threonine residues in specific proteins.

Stereoisomers of threonine. (Palyi G., 2019)

Threonine pKa

The pKa values for threonine correspond to its α-carboxyl group (pKa = 2.1) and α-amino group (pKa ≈ 9.1). At physiological pH (~7.4), the carboxyl group is deprotonated (–COO^⁻), the amino group is protonated (–NH3⁺), and the side chain (–CH(OH)CH₃) remains neutral due to the absence of an ionizable group. Consequently, threonine resides in a zwitterionic state at physiological pH, where the positive and negative charges neutralize each other, resulting in a net charge of zero for the molecule.

Threonine solubility

It is anticipated that huge three-dimensional crystals would form when a solvent with a smaller dipole moment and the optimal solubility is chosen. According to these points of view, experiments on the solubility of L-threonine in several solvents, including water, methanol, ethanol, and combinations of these, have been conducted. A single crystal of L-threonine was most effectively crystallized in deionized water due to the fact that this polar amino acid has a dipole moment almost identical to water's.

Solubility of L-threonine in water solvent. (Ramesh Kumar G., et al., 2009)

Threonine amino acids at Creative Peptides

Difference between serine and threonine

Serine and threonine are polar, hydrophilic amino acids characterized by hydroxyl groups in their side chains, enabling them to establish hydrogen bonds and engage in phosphorylation. The primary distinction between them is that serine possesses a smaller hydroxymethyl group (–CH₂OH), whereas threonine contains a larger hydroxyethyl group (–CH(OH)CH₃) with an extra methyl group (-CH₃). The additional methyl group renders threonine marginally more polar and introduces increased steric hindrance in proteins. Threonine is an essential amino acid that must be acquired through dietary sources, whereas serine is non-essential and can be produced from glycine within the body. Notwithstanding these distinctions, both are crucial to protein metabolism and functionality.

Molecular structures of amino acids. (Zhang D Q., et al., 2008)

Threonine metabolism

Threonine is utilized as a substrate for the production of mucin, which is a protein. Additionally, Thr has the ability to cross over into the catabolic pathway, where it can be converted into a wide range of essential metabolites (glycine, acetyl CoA, and pyruvate) that are essential to the metabolism of the host. There are three distinct metabolic circuits that threonine goes through. The glycine-independent or glycine-dependent pathway is that which is followed by the catabolism of Thr. Thr is converted to α-ketobutyric acid and ammonia by liver threonine dehydratase (STDH) through the glycine-independent pathway, as demonstrated by a study. Following this, the ketobutyrate is decarboxylated to generate propionyl-CoA. As a result of this process, the ketobutyrate is responsible for the formation of propionyl-CoA. When it comes to the pancreas and liver, respectively, threonine dehydrogenase (TDH) and somatotransferase (STDH) are abundantly expressed. On the other hand, threonine aldolase is highly expressed in the prostate rather than the liver, where it has a modest degree of enzymatic activity. As part of the glycine-dependent process, TDH and threonine aldolase are involved. In the breakdown of Thr, TDH is an essential enzyme. The enzyme 2-amino-3-oxobutyrate CoA ligase (GCAT) is responsible for the conversion of Thr to 2-amino-3-oxybutyrate, which is an unstable intermediate that is subsequently reduced to acetyl-CoA and glycine. After that, acetyl-CoA is introduced into the tricarboxylic acid (TCA) cycle, within which it contributes to the production of energy. Further, threonine aldolase is responsible for the conversion of Thr into glycine and acetaldehyde. The physiologic state can have an effect on the pathway of Thr metabolism, which is something that should be taken into consideration. It is only the STDH pathway that is responsible for the degradation of Thr in newborns; however, in adults, the TDH pathway is responsible for the catabolism of 7–10% of the total Thr that is present. Glycine is required in greater quantities by newborns than by adults, which is the reason for this discrepancy.

The metabolic pathway of Threonine. (Tang Q., et al., 2021)

Function of threonine

(1) Protein synthesis

In epithelial tissues, like mucins, threonine is necessary for the production of Thr-rich proteins. Animals produce less gut mucosal protein and mucins when their food Thr concentration is low or high, demonstrating how sensitive mucin synthesis is to dietary Thr concentration. Thr is beneficial for protein synthesis in various tissues, including skeletal muscles, in addition to intestinal mucus. Research on several aquatic animals and cattle has shown that Thr can stimulate growth and improve skeletal muscle protein synthesis. In addition to being a building block for proteins, threonine can control the process by acting as a signaling molecule. Muscle growth and repair depend on insulin-like growth factor I (IGF-I), which acts as an upstream activator of mammalian target of rapamycin (mTOR) in skeletal muscle. The PI3K/AKT pathway regulates protein synthesis via ribosomal S6 kinase (S6K) and the eukaryotic translation initiation factor 4E-binding protein 1 (4E-BP1), of which the mTOR is a downstream component. The activation of the PI3K/AKT/TOR signaling cascade via IGF-I in fish was found to be enhanced muscle protein synthesis in response to dietary Thr, according to a recent publication. Thr is broken down by TDH into glycine, which is a precursor to glycine. Through suppressing proteolysis and activating mTORC1 in a PI3K/Akt-dependent way, glycine regulates protein synthesis.

(2) Regulate lipid metabolism

The metabolism of lipids is closely related to essential amino acids; specifically, methionine, leucine, and isoleucine can influence the deposition of lipids. Thr also plays a role in controlling the accumulation and metabolism of lipids. In contrast to Thr depletion, which may cause hepatic triglyceride buildup, Thr supplementation improves hepatic lipid metabolism. A study conducted on Pekin ducks revealed that a lack of Thr in their diet led to an upregulation of genes associated to triglyceride and fatty acid synthesis, including 3-oxoacyl-ACP synthase (OXSM), very-long-chain fatty acids protein 7 (ELOVL7), and long-chain fatty acyl-CoA ligase (ACSBG2). The following genes were found to be downregulated in Pekin ducks when Thr was absent: acyl-CoA dehydrogenase, acyl-CoA dehydrogenase family member 11, cytochrome P450 (CYP4B), apolipoprotein D (APOD), and microsomal triglyceride transfer protein (MTTP), all of which are involved in fatty acid oxidation. Maintaining a healthy balance of lipids is dictated by the constant regulation of lipogenesis and lipolysis. Lipogenesis is regulated by adipogenic transcription factors, such as peroxisome proliferator-activated receptors (PPARs), which help preadipocytes to transform into mature adipocytes. PPARγ modulates gene networks related to glucose homeostasis, including increasing the expression of glucose transporter type 4 (Glut4) and c-Cbl-associated protein (CAP), and is essential for adipocyte development, lipid metabolism, and insulin sensitivity. By modulating the PPARγ signaling pathway, threonine supplementation in a high-fat diet has the potential to reduce lipid accumulation. Brown adipose tissue expresses the thermogenetic-related gene uncoupling protein-1 (UCP1), which can dissipate energy as heat in the battle against obesity. Brown adipose tissue UCP1 expression was downregulated in obese mice, but it was upregulated again when Thr was added to a high-fat diet. Cardiovascular disease can be exacerbated by alterations in lipid metabolism. Some lipid metabolic diseases may be positively modulated by threonine. There is a clear negative correlation between Thr and levels of small dense low-density lipoprotein cholesterol (sdLDL-C) and triglycerides (TG), and one study found that elevated plasma Thr levels are associated with a reduced risk of atherogenic lipid profiles in adults.

(3) Maintain stem cell function

Preimplantation mammalian blastocysts develop two layers of cells, the trophectoderm and the inner cell mass (ICM), a few days after the merging of sperm and egg cells. Embryos make touch with the uterine epithelium during implantation through the trophectoderm, a single layer of cells on their surface. From the interstitial lining (ICM) emerge all the various tissues of mammals. The ICM is both a source of and a model for embryonic stem (ES) cells. In order to keep ES cells functioning, threonine is essential. Neither proliferation nor undifferentiation of murine (mES) and likely bovine ES cells is possible in the absence of Thr in their culture media. Threonine dehydrogenase (TDH) converts Thr to glycine and acetyl CoA in these endosomal cells. Posttranslational and posttranscriptional effectors, in addition to its own genes, modulate TDH activity. In order to control epigenetic changes, nucleotide biosynthesis, and mitochondrial free energy conversions, these cells rely on the glycine and acetyl CoA produced by TDH, which are essential precursors. Threonine enhances mTOR-mediated signaling in mES cells by promoting cMyc expression. Damage to lipid rafts that contain Thr transporter(s) prevents this Thr-initiated signaling from taking place. It is believed that 3-hydroxynorvaline (3-HNV), a Thr analog, inhibits mES cell proliferation by blocking Thr degradation and competing with TDH for Thr transport. Similarly, this metabolic signaling may aid blastocysts in retaining pluripotency and proliferation within their inner cell membranes (ICMs). As mentioned before, Thr builds up in mouse blastocysts as they get closer to implantation in the uterus, which is different from other amino acids.

What is the threonine used for?

(1) Improve intestinal health

The body makes use of the dietary threonine that is absorbed by the intestines for either the synthesis of proteins or the oxidation of it in order to produce energy. In terms of the use and metabolism of amino acids, the gut is the most important location. Threonine is not only involved in the production of mucosal proteins, but it is also broken down by luminal bacteria that are found in the intestines. Although the use of threonine for oxidation in the intestines of pigs only accounts for 2–9% of the total Thr, the use of threonine for the synthesis of mucosal proteins accounts for 71% of the overall Thr utilization. The peptide backbone of mucin contains a significant amount of Thr, which is responsible for thirty percent of the total amino acids found in this protein. Because of the limited availability of Thr in the diet, muscle growth and other activities of the tissues are constrained, which comes at the expense of maintaining mucin formation. In the gut, certain commensal bacteria, namely species of Clostridium, have the ability to breakdown Thr. This process leads to the generation of volatile fatty acids, including acetic acid, propionic acid, and butyric acid. These acids are essential for the maintenance of intestinal function and the modulation of immune responses.

Gut function encompasses a wide range of functions, including the digestion and absorption of nutrients, as well as the immune system's defense against viruses and toxins. To ensure that the intestinal function of nutrient uptake and absorption is carried out properly, the integrity of the intestinal structure must be maintained. Additionally, the intestinal mucosa is able to absorb and make use of an adequate amount of Thr, which helps to ensure that the mucosa continues to be intact. A beneficial effect on villus height, crypt depth, epithelial thickness, and the quantity of goblet cells is observed in the small intestine of broilers when dietary Thr supplementation is administered. By reducing villous area, inducing villous atrophy, and increasing the apoptotic rate of intestinal epithelial cells in weaned pigs, dietary Thr that is either insufficient or excessive may be responsible for these effects. The higher villus helps to enhance the amount of surface area that is available for the consumption of nutrients. Additionally, the Thr need for amylase synthesis is exceptionally high, contributing for around 11% of the total protein. According to the findings of a number of studies, the absence of Thr in the diet leads to a decrease in the production of digestive enzymes as well as the activity of brush border enzymes in the digestive tract of animals. With this in mind, it is plausible that Thr could potentially contribute to the enhancement of intestine digesting and absorptive ability.

(2) Regulate gut microbiota

The bacteria in the gut are more favorable when dietary Thr is consumed. In broiler hens, a high dietary Thr reduced the number of colonies of Salmonella and Escherichia coli (E. coli), while simultaneously increased the number of Lactobacillus. In addition, research has demonstrated that administering dietary Thr to animals in times of stress helps to restore the balance of the microbiota in their guts. The microbial diversity was regained and the quantity of potentially helpful bacteria was dramatically increased in laying hens that were fed a diet that included a low amount of crude protein and was supplemented with Thr at a concentration of 0.3%. The prevalence of beneficial communities of Enterobacteria, Lactobacillus, Bacteroides, and Enterococci was raised in rats that were challenged with dextran sodium sulfate when they were given an amino acid cocktail that contained L-Thr. One of the possible causes for this action is that Thr stimulates mucin secretion. This is due to the fact that mucins are unable to be digested in the small intestine, and as a result, they make their way to the large intestine, where they act as substrates for saccharolytic bacteria. According to the findings of Koo's team, the addition of Thr at a level that is fifteen percent greater than the recommendation made by the NRC can interact with the content of feed to change the fermentation of gut microbes. The studies also suggest that Thr has favorable effects on the microbiota of the intestinal tract, which may be related to the development of immunoglobulin and mucin as a result of the addition of Thr.

(3) Improve barrier function

The mucus layer covering the intestinal epithelium is essential for intestinal barrier function. The mucus layer is a physical barrier composed of mucins, immunoglobulins, salts, antibacterial substances, and other components, released by various glands. The mucus layer acts as the primary barrier against harm from digestive enzymes, bacteria, and pathogens. Mucin-2 (MUC2) is the principal component of intestinal mucus, predominantly synthesized by goblet cells. MUC2 is a heavily glycosylated glycoprotein characterized by a core protein backbone abundant in threonine and serine, coupled to several O-linked oligosaccharide side chains, resulting in significant resistance to proteolysis. Threonine is the primary regulator of intestinal mucin secretion. In comparison to sufficient Thr (0.89%), both deficiency and surplus of Thr (0.37% and 1.11%, respectively) markedly influence the quantity and composition of intestinal mucin in piglets. Nonetheless, the mechanism by which high dietary threonine diminishes the synthesis of intestinal mucosal proteins and mucins remains unclear. A plausible explanation is that neutral amino acids, notably branched-chain amino acids, utilize the same transport channels as threonine, thereby constraining its intestinal absorption and subsequently restricting protein synthesis. This potential explanation necessitates additional investigation. Growing data indicates that goblet cell development and mucin synthesis are responsive to dietary threonine levels. Increased dietary Thr supplementation beyond growth needs resulted in improved goblet cell density and MUC2 mRNA expression in the intestine. A study suggested that dietary threonine may influence goblet cell development by regulating the expression of genes involved in the Notch-Hes1-Math1 pathway, hence enhancing MUC2 synthesis. The Notch signaling system is crucial for the differentiation of intestinal cells. Notch signaling stimulates the Hes1 transcription factor and suppresses the bHLH transcription factor Math1 (the human equivalent is Hath1). Activation of Hath1 induces MUC2 expression by binding to Eboxes located on the MUC2 promoter. MUC2 serves both as a physical barrier and as a regulator of immunological responses, interacting with the epithelium, microbiota, and the host immune system to preserve intestinal homeostasis. Furthermore, the influence of Thr on mucin production is associated with age and physiological condition. Research indicated that 45% of the dietary threonine requirement is necessary for the maintenance of intestinal mucosa in three-day-old pigs. Studies on animals indicate that in the presence of disease and stress circumstances, including ileitis, sepsis, inflammation, and intrauterine development retardation, the intestinal demand for Thr significantly increases to enhance mucin synthesis. This indicates that intestinal inflammation elevates mucus production to safeguard the gut. In conclusion, dietary Thr supplementation enhances MUC2 synthesis, hence augmenting MUC2's impact on gut barrier integrity, immunological function, and the maintenance of intestinal homeostasis.

Moreover, numerous recent studies have concentrated on the impact of the interplay between dietary threonine and dietary fiber on intestinal barrier integrity. Dietary fiber enhances barrier function by increasing mucus secretion and mucin production. Nevertheless, elevated dietary fiber enhances the depletion of endogenous amino acids, hence necessitating a higher requirement for threonine to sustain mucin synthesis. The impact of Thr on intestinal barrier proteins is contingent upon dietary fiber levels. The expression of barrier protein increased with elevated Thr levels in pigs consuming a low-fiber diet, rather than a high-fiber diet.

The effects of Thr levels on intestinal barrier function. (Sahoo D P., et al., 2021)

(4) Prevent fat deposition

Obesity is a major contributor to the development of numerous diseases. Excess fat and serum triglycerides can be diminished in obese mice by supplementing with threonine. It is not yet known, however, if threonine can prevent obesity in mice that have not been previously fed a high-fat diet. For 15 weeks, mice were given either a chow diet (CD) or a high-fat diet (HFD), with or without threonine supplementation (3.0% in drinking water). When compared to mice fed a HFD, those given chronic threonine supplementation had lower levels of total cholesterol, serum low-density lipoprotein cholesterol, epididymal white adipose tissue weight, and body weight. Sterol regulatory element-binding protein 1c and fatty acid synthase gene expressions were up-regulated in the epididymal adipose tissue, whereas hormone sensitive lipase, adiponectin, and fibroblast growth factor 21 gene expressions were down-regulated. In HFD-fed animals, threonine supplementation up-regulated gene expressions of sirtuin1, adenosine monophosphate-activated protein kinase, and peroxisome proliferator activated receptor γ co-activator 1α in the liver tissue. Based on these findings, threonine supplementation may be a useful tool in the fight against diet-induced obesity as it reduces fat mass and enhances lipid metabolism when used over the long term.

Threonine (Thr) supplementation alleviated fat deposition. (Chen J., et al., 2022)

FAQ

Is threonine hydrophobic or hydrophilic?

Threonine exhibits hydrophilicity. This is attributable to its polar side chain, which has a hydroxyl group (–OH) linked to a hydroxyethyl group (–CH(OH)CH₃). The hydroxyl group can establish hydrogen bonds with water molecules, rendering threonine polar and water-soluble. Consequently, threonine is generally located on the outside of proteins, engaging with the aqueous milieu. Conversely, hydrophobic amino acids possess nonpolar side chains that exhibit poor interaction with water. Threonine's capacity to create hydrogen bonds and its polar characteristics categorize it as hydrophilic.

Is threonine acidic or basic?

No, it is a neutral amino acid. The side chain, comprising a hydroxyethyl group (–CH(OH)CH₃), is neutral and does not undergo ionization. The α-carboxyl group (–COOH) can dissociate a proton to provide –COO^⁻ at normal pH (~7.4); nonetheless, this characteristic is typical of all amino acids and does not render threonine acidic. The α-amino group (–NH2) is protonated to provide –NH3⁺. At physiological pH, threonine adopts a zwitterionic configuration, exhibiting no net charge, hence underscoring its neutrality. Consequently, threonine is categorized as neutral instead than acidic.

Is threonine polar?

Indeed, threonine is a polar amino acid. The side chain includes a hydroxyl group (–OH), enabling it to establish hydrogen bonds with water and other polar molecules. The hydroxyethyl group (–CH(OH)CH₃) in the side chain enhances its polarity, rendering threonine a hydrophilic amino acid. The polarity of threonine facilitates its interaction with aquatic environments, and it is frequently located on the surface of proteins, enabling the formation of hydrogen bonds. Consequently, threonine is categorized as a polar amino acid.

Is threonine an essential amino acid?

Indeed, threonine is classified as an essential amino acid since it cannot be produced by the human body and must be acquired through dietary sources. Threonine is essential for protein synthesis, metabolism, and enzymatic activity, and it contributes to the creation of crucial compounds such as collagen and elastin. Threonine, being an essential amino acid, must be ingested from dietary sources such as meat, dairy, eggs, beans, and lentils, as the body is incapable of synthesizing it.

References

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