Pathways

3-Methylhistidine, also known as 3-MHis, 3MH, pi-methylhistidine or pros-methylhistidine, belongs to the class of organic compounds known as histidine and derivatives. 3MH is also classified as a methylamino acid. Methylamino acids are primarily proteogenic amino acids (found in proteins) which have been methylated (in situ) on their side chains by various methyltransferase enzymes. Histidine can be methylated at either the N1 or N3 position of its imidazole ring, yielding the isomers 1-methylhistidine (1MH; also referred to as tau-methylhistidine, according to IUPAC) or 3-methylhistidine (3MH; pi-methylhistidine, according to IUPAC), respectively. There is considerable confusion with regard to the nomenclature of the methylated nitrogen atoms on the imidazole ring of histidine in histidine-containing proteins (such as actin and myosin) as well as histidine-containing peptides (such as anserine and ophidine/balenine). In particular, older literature (mostly prior to the year 2000) as well as most biochemists and nutrition scientists incorrectly number the imidazole nitrogen atom most proximal to the side chain beta-carbon as 1 or N1, while organic chemists correctly designate it as 3 or N3. As a result, biochemists and nutrition scientists historically designated anserine (Npi-methylated) as beta-alanyl-N1-methylhistidine (or beta-alanyl-1-methylhistidine), whereas according to standard IUPAC nomenclature, anserine is correctly named as beta-alanyl-N3-methylhistidine. As a result, for several decades, many papers incorrectly identified 1MH as a specific marker for dietary consumption or various pathophysiological effects when they really are referring to 3MH – and vice versa. 3MH can only be generated from histidine residues through the action of methyltransferases as a protein post-translational modification event. Histidine methylation on the 3- or pi site of histidine-containing proteins is mediated by only one known enzyme – METTL9. Recent discoveries have shown that 3MH is produced in essentially all vertebrates via the methyltransferase enzyme known as METTL9. METTL9 is a broad-specificity S-adenosylmethionine-mediated methyltransferase that mediates the formation of the majority of 3MH present in mammalian and other vertebrate proteomes. Because of its abundance in some muscle-related proteins but especially because of the high abundance of anserine found in poultry and fish, 3MH has been found to be a good biomarker for the consumption of meat. Dietary studies have shown that general poultry consumption (p-trend = 0.0006) and especially chicken consumption (p-trend = 0.0003) are associated with increased levels of 3MH in human plasma. 3‐MH is synthesized only in the muscle by the methylation of one histidine residue in actin and in varying amounts in myosin depending on the type of muscle. Thus, muscle protein degradation is the only endogenous source of 3‐MH in human plasma. 3‐MH might be a helpful biomarker in the assessment of muscle protein turnover, which is important in the diagnosis of frailty and sarcopenia.

4-Hydroxyhippuric, also known as 4-hydroxybenzoylglycine or 4-hydroxyhippate acid, is a metabolite of hippuric acid, and a uremic toxin. Benzoic acid is present in many fruits, such as apricots, prunes, and berries; many vegetables such as mushrooms (fungus), snap peas, cucumbers, and radishes; spices such as cinnamon, cloves, and allspice; nuts such as​ cashews, almonds, pistachios; and dairy products such as yogurt, milk, and cheese. Benzoic acid can also be synthesized by gut microbes through phenylalanine, however the exact mechanisms of synthesis in microbes is not well studied. Benzoic acid is transported out of the intestine via a monocarboxylate transporter into the blood. Then it is transported into the liver via another monocarboxylate transporter. In the mitochondira of the liver benzoic acid is catalyzed by the enzyme acyl-coenzyme A synthetase ACSM2A, mitochondrial, with ATP and coenzyme A into the metabolite Benzoyl-CoA. Benzoyl-CoA is then catalyzed by the enzyme glycine N-acyltransferase and a glycine, which produces hippuric acid. Hippuric acid leaves the mitochondria and is metabolized in the membrane of the endoplasmic reticulum by the enzyme cytochrome P450 1A2 to produce 4-Hydroxyhippuric. 4-Hydroxyhippuric is transported into the blood by a monocarboxylate transporter. 4-Hydroxyhippuric inhibits the Ca2+-ATPase on the plasma membrane of erythrocytes. This leads to apoptosis of the erythrocytes. It also induces free radical production in the renal proximal tubular cell line.

p-Hydroxyphenylacetic acid, also known as 4-hydroxybenzeneacetate, is classified as a member of the 1-hydroxy-2-unsubstituted benzenoids. 1-Hydroxy-2-unsubstituted benzenoids are phenols that are unsubstituted at the 2-position. p-Hydroxyphenylacetic acid is considered to be slightly soluble (in water) and acidic. p-Hydroxyphenylacetic acid can be synthesized from acetic acid. It is also a parent compound for other transformation products, including but not limited to, methyl 2-(4-hydroxyphenyl)acetate, ixerochinolide, and lactucopicrin 15-oxalate. p-Hydroxyphenylacetic acid can be found in numerous foods such as olives, cocoa beans, oats, and mushrooms. p-Hydroxyphenylacetic acid can be found throughout all human tissues and in all biofluids. Within a cell, p-hydroxyphenylacetic acid is primarily located in the cytoplasm and in the extracellular space. p-Hydroxyphenylacetic acid is also a microbial metabolite produced by Acinetobacter, Clostridium, Klebsiella, Pseudomonas, and Proteus. Higher levels of this metabolite are associated with an overgrowth of small intestinal bacteria from Clostridia species including C. difficile, C. stricklandii, C. lituseburense, C. subterminale, C. putrefaciens, and C. propionicum. l-tyrosine, derived from diet and endogenous proteins and peptides, can be converted to phenol and 4-hydroxyphenylpyruvate. Tyrosine phenol-lyase (EC 4.1.99.2.), previously named β–tyrosinase, is responsible for the reversible deamination of l-tyrosine, requiring pyridoxyl phosphate as a cofactor, into phenol ammonia and pyruvate. This reaction is also reversible by the same enzyme using l-serine and phenol as substrates. In addition, the reversible reaction of l-tyrosine with 2-oxoglutarate in 4-hydroxyphenylpyruvate and L-glutamate is catalysed by tyrosine transaminase (EC 2.6.1.5.) or by aromatic-amino-acid transaminase (EC 2.6.1.57.). To a small extent, 4-hydroxyphenylpyruvate and ammonia can also be formed by the enzyme phenylalanine dehydrogenase (EC 1.4.1.20.) from l-tyrosine. -Hydroxyphenylpyruvate is the precursor of 4-hydroxyphenylacetate, catalysed by p-hydroxyphenylpyruvate oxidase (EC 1.2.3.13.).

Uric acid is formed from purine catabolism. Purines can be made endogenously in the body or can be obtained exogenously from foods such as red meat. The purines are guanine and adenine. These undergo metabolism in the liver to form uric acid. Adenine forms adenosine through the enzyme purine nucleoside phosphorylase. Adenosine then reacts with water to form inosine and ammonia using the enzyme adenosine deaminase. Inosine goes on to form hypoxanthine through the enzyme purine nucleoside phosphorylase. Xanthine is formed from hypoxanthine using the enzyme xanthine dehydrogenase/ oxidase. Xanthine can also be formed from the purine guanine via guanine deaminase. Uric acid is produced from xanthine in the presence of xanthine dehydrogenase/ oxidase. Uric acid is the final oxidation product of purine (adenine and guanine) metabolism in humans and higher primates, and is removed from renal and gastrointestinal routes. In lower animals such as rats and mice, the enzyme uricase (urate oxidase) further oxidizes uric acid to allantoin for more efficient removal from the urine. Humans and higher primates lack a functional uricase gene. The enzyme urate oxidase (UO), uricase or factor-independent urate hydroxylase, absent in humans, catalyzes the oxidation of uric acid to 5-hydroxyisourate. Urate oxidase is the first enzyme in a pathway of three enzymes to convert uric acid to S-(+)-allantoin. After uric acid is converted to 5-hydroxyisourate by urate oxidase, 5-hydroxyisourate (HIU) is converted to 2-oxo-4-hydroxy-4-carboxy-5-ureidoimidazoline (OHCU) by HIU hydrolase, and then to S-(+)-allantoin by 2-oxo-4-hydroxy-4-carboxy-5-ureidoimidazoline decarboxylase (OHCU decarboxylase). Without HIU hydrolase and OHCU decarboxylase, HIU will spontaneously decompose into racemic allantoin. Urate oxidase is found in nearly all organisms, from bacteria to mammals, but is inactive in humans and several other great apes, having been lost in primate evolution.

Uric acid is formed from purine catabolism. Purines can be made endogenously in the body or can be obtained exogenously from foods such as red meat. The purines are guanine and adenine. These undergo metabolism in the liver to form uric acid. Adenine forms adenosine through the enzyme purine nucleoside phosphorylase. Adenosine then reacts with water to form inosine and ammonia using the enzyme adenosine deaminase. Inosine goes on to form hypoxanthine through the enzyme purine nucleoside phosphorylase. Xanthine is formed from hypoxanthine using the enzyme xanthine dehydrogenase/ oxidase. Xanthine can also be formed from the purine guanine via guanine deaminase. Uric acid is produced from xanthine in the presence of xanthine dehydrogenase/ oxidase. Uric acid is the final oxidation product of purine (adenine and guanine) metabolism in humans and higher primates, and is removed from renal and gastrointestinal routes. In lower animals such as rats and mice, the enzyme uricase (urate oxidase) further oxidizes uric acid to allantoin for more efficient removal from the urine. Humans and higher primates lack a functional uricase gene. The enzyme urate oxidase (UO), uricase or factor-independent urate hydroxylase, absent in humans, catalyzes the oxidation of uric acid to 5-hydroxyisourate. Urate oxidase is the first enzyme in a pathway of three enzymes to convert uric acid to S-(+)-allantoin. After uric acid is converted to 5-hydroxyisourate by urate oxidase, 5-hydroxyisourate (HIU) is converted to 2-oxo-4-hydroxy-4-carboxy-5-ureidoimidazoline (OHCU) by HIU hydrolase, and then to S-(+)-allantoin by 2-oxo-4-hydroxy-4-carboxy-5-ureidoimidazoline decarboxylase (OHCU decarboxylase). Without HIU hydrolase and OHCU decarboxylase, HIU will spontaneously decompose into racemic allantoin. Urate oxidase is found in nearly all organisms, from bacteria to mammals, but is inactive in humans and several other great apes, having been lost in primate evolution.

Uric acid is formed from purine catabolism. Purines can be made endogenously in the body or can be obtained exogenously from foods such as red meat. The purines are guanine and adenine. These undergo metabolism in the liver to form uric acid. Adenine forms adenosine through the enzyme purine nucleoside phosphorylase. Adenosine then reacts with water to form inosine and ammonia using the enzyme adenosine deaminase. Inosine goes on to form hypoxanthine through the enzyme purine nucleoside phosphorylase. Xanthine is formed from hypoxanthine using the enzyme xanthine dehydrogenase/ oxidase. Xanthine can also be formed from the purine guanine via guanine deaminase. Uric acid is produced from xanthine in the presence of xanthine dehydrogenase/ oxidase. Uric acid is the final oxidation product of purine (adenine and guanine) metabolism in humans and higher primates, and is removed from renal and gastrointestinal routes. In lower animals such as rats and mice, the enzyme uricase (urate oxidase) further oxidizes uric acid to allantoin for more efficient removal from the urine. Humans and higher primates lack a functional uricase gene. The enzyme urate oxidase (UO), uricase or factor-independent urate hydroxylase, absent in humans, catalyzes the oxidation of uric acid to 5-hydroxyisourate. Urate oxidase is the first enzyme in a pathway of three enzymes to convert uric acid to S-(+)-allantoin. After uric acid is converted to 5-hydroxyisourate by urate oxidase, 5-hydroxyisourate (HIU) is converted to 2-oxo-4-hydroxy-4-carboxy-5-ureidoimidazoline (OHCU) by HIU hydrolase, and then to S-(+)-allantoin by 2-oxo-4-hydroxy-4-carboxy-5-ureidoimidazoline decarboxylase (OHCU decarboxylase). Without HIU hydrolase and OHCU decarboxylase, HIU will spontaneously decompose into racemic allantoin. Urate oxidase is found in nearly all organisms, from bacteria to mammals, but is inactive in humans and several other great apes, having been lost in primate evolution.

Androsterone sulfate (Andros-S) is the most abundant 5-alpha-reduced androgen metabolite in serum. Androsterone sulfate is clinically recognized as one of the major androgen metabolites found in urine. It is a cognate substrate for human dehydroepiandrosterone sulfotransferase, which catalyzes the transfer of the sulfonate group from 3'-phosphoadenosine-5'-phosphosulfate to dehydroepiandrosterone (DHEA). Androsterone sulfate has been identified in the human placenta. Testosterone is the primary male sex hormone and anabolic steroid in males. Like other steroid hormones, testosterone is derived from cholesterol. The largest amounts of testosterone (>95%) are produced by the testes in men, while the adrenal glands account for most of the remainder. Both testosterone and 5α-DHT are metabolized mainly in the liver. Approximately 50% of testosterone is metabolized via conjugation into testosterone glucuronide and to a lesser extent testosterone sulfate by glucuronosyltransferases and sulfotransferases, respectively. In the hepatic 17-ketosteroid pathway of testosterone metabolism, testosterone is converted in the liver by 5α-reductase and 5β-reductase into 5α-DHT and the inactive 5β-DHT, respectively. Then, 5α-DHT and 5β-DHT are converted by 3α-HSD into 3α-androstanediol and 3α-etiocholanediol, respectively. Subsequently, 3α-androstanediol and 3α-etiocholanediol are converted by 17β-HSD into androsterone and etiocholanolone, which is followed by their conjugation and excretion. Androsterone has generally been considered to be an inactive metabolite of testosterone, which when conjugated by glucuronidation and sulfation allows testosterone to be removed from the body. Androsterone is sulfated into androsterone sulfate. Androsterone sulfate, also known as 3α-hydroxy-5α-androstan-17-one 3α-sulfate, is an endogenous, naturally occurring steroid and one of the major urinary metabolites of androgens. It is a steroid sulfate which is formed from sulfation of androsterone by the steroid sulfotransferase SULT2A1 and can be desulfated back into androsterone by steroid sulfatase.

beta-Alanine is the only naturally occurring beta-amino acid - an amino acid in which the amino group is at the beta-position from the carboxylate group. It is formed in vivo by the degradation of dihydrouracil and carnosine. It is a component of the naturally occurring peptides carnosine and anserine and also of pantothenic acid (vitamin B-5), which itself is a component of coenzyme A. Under normal conditions, beta-alanine is metabolized into acetic acid. beta-Alanine can undergo a transanimation reaction with pyruvate to form malonate-semialdehyde and L-alanine. The malonate semialdehyde can then be converted into malonate via malonate-semialdehyde dehydrogenase. Malonate is then converted into malonyl-CoA and enter fatty acid biosynthesis. Since neuronal uptake and neuronal receptor sensitivity to beta-alanine have been demonstrated, beta-alanine may act as a false transmitter replacing gamma-aminobutyric acid. When present in sufficiently high levels, beta-alanine can act as a neurotoxin, a mitochondrial toxin, and a metabotoxin. A neurotoxin is a compound that damages the brain or nerve tissue. A mitochondrial toxin is a compound that damages mitochondria and reduces cellular respiration as well as oxidative phosphorylation. A metabotoxin is an endogenously produced metabolite that causes adverse health effects at chronically high levels. Chronically high levels of beta-alanine are associated with at least three inborn errors of metabolism, including GABA-transaminase deficiency, hyper-beta-alaninemia, and methylmalonate semialdehyde dehydrogenase deficiency. beta-Alanine is a central nervous system (CNS) depressant and is an inhibitor of GABA transaminase. The associated inhibition of GABA transaminase and displacement of GABA from CNS binding sites can also lead to GABAuria (high levels of GABA in the urine) and convulsions. In addition to its neurotoxicity, beta-alanine reduces cellular levels of taurine, which are required for normal respiratory chain function. Cellular taurine depletion is known to reduce respiratory function and elevate mitochondrial superoxide generation, which damages mitochondria and increases oxidative stress. Individuals suffering from mitochondrial defects or mitochondrial toxicity typically develop neurotoxicity, hypotonia, respiratory distress, and cardiac failure. beta-Alanine is a biomarker for the consumption of meat, especially red meat. A main pathway of beta-Alanine biosynthesis is degradation of beta-Alanyl-(L)-histidine (carnosine). Carnosine N-methyltransferase (EC 2.1.1.22) converts beta-Alanyl-(L)-histidine to Anserine using S-Adenosyl-L-methionine as methyl donor. Then Anserine is hydrolyzed to beta-Alanine by Carnosine dipeptidase 1 (metallopeptidase M20 family) CPGL2. Beta-Alanyl-(L)-histidine may also be hydrolyzed by CPGL2 to beta-Alanine and (L)-Histidine. Dietary sources of carnosine include meats, eggs, and dairy products, with meat being the primary source.

Choline is obtained from foods like poultry, meat, fish, dairy and eggs. In the intestine, choline is metabolized to other products such as acetylcholine, betaine, phosphatidylcholine and trimethylamine (TMA) by the gut microbiota. The toxicity of choline is due to the formation of TMA and its metabolism to Trimethylamine N-oxide (TMAO) in the liver. TMA is created from choline via the enzyme choline trimethylamine lyase in the intestinal microbe then TMA then enters the bloodstream and is transported to the liver where Dimethylaniline monooxygenase [N-oxide-forming] 3 converts TMA to TMAO. TMAO has negative effects on organs such as the heart, kidney and vascular system by contributing to cardiovascular disease, atherosclerosis, endothelial dysfunction and kidney disease.

Citrulline, also known as Cit or δ-ureidonorvaline, belongs to the class of organic compounds known as l-alpha-amino acids. These are alpha amino acids which have the L-configuration of the alpha-carbon atom. Citrulline has the formula H2NC(O)NH(CH2)3CH(NH2)CO2H. Citrulline exists in all living species, ranging from bacteria to humans. Within humans, citrulline participates in a number of enzymatic reactions. In particular, citrulline can be biosynthesized from carbamoyl phosphate and ornithine which is catalyzed by the enzyme ornithine carbamoyltransferase. In addition, citrulline and L-aspartic acid can be converted into argininosuccinic acid through the action of the enzyme argininosuccinate synthase. In humans, citrulline is involved in the metabolic disorder called argininemia. Citrulline has also been found to be associated with several diseases such as ulcerative colitis, rheumatoid arthritis, and citrullinemia type II. Citrulline has also been linked to several inborn metabolic disorders including argininosuccinic aciduria and fumarase deficiency. Outside of the human body, citrulline is found, on average, in the highest concentration in a few different foods such as wheats, oats, and cucumbers and in a lower concentration in swiss chards, yellow wax beans, and potato. Citrulline has also been detected, but not quantified in several different foods, such as epazotes, lotus, common buckwheats, strawberry guava, and italian sweet red peppers. Citrulline is a potentially toxic compound. Proteins that normally contain citrulline residues include myelin basic protein (MBP), filaggrin, and several histone proteins, whereas other proteins, such as fibrin and vimentin are susceptible to citrullination during cell death and tissue inflammation. Citrulline is also produced as a byproduct of the enzymatic production of nitric oxide from the amino acid arginine, catalyzed by nitric oxide synthase. It is also produced from arginine as a byproduct of the reaction catalyzed by NOS family (NOS; EC1.14.13.39). Ornithine transcarbamoylase (OTC) or OCT (EC 2.1. 3.3) is an enzyme that catalyzes the reaction of citrulline formation from l-ornithine and carbamoyl phosphate. In mammals it is almost exclusively located in the mitochondria of hepatocytes and is part of the urea cycle. It catalyzes the second step of the urea cycle, the condensation of carbamoyl phosphate with L-ornithine to form L-citrulline (carbamoyl phosphate + L-ornithine = H+ + L-citrulline + phosphate).