Tetrahydrofolate (THF) or tetrahydrofolic acid is a derivative of Vitamin B9 (folic acid or pteroyl-L-glutamic acid), a water-soluble vitamin that serves as a coenzyme for metabolic reactions involving amino acids and nucleic acids. It participates in important single-carbon transfer reactions often referred to as one-carbon metabolism and in the synthesis of several amino acids such as serine and methionine, purines and thymine, a pyrimidine. The single carbon transfer reactions are important in metabolism and regulation of gene expression.
Chemically folic acid consists of three structural components- para-aminobenzoic acid (PABA), a bicyclic pteridine ring, and glutamic acid. The active form of folic acid is known as tetrahydrofolic acid or tetrahydrofolate (THF or FH4), which is a result of the addition of four hydrogen atoms to 5,6,7 and 8 position in the pteridine ring.
Tetrahydrofolate is involved in single carbon transfer that is bound to positions N5, N10, or both. Serine, glycine, histidine, and tryptophan are the carbon sources for the one-carbon pool, serine being the major source. They donate the single carbon to THF in various oxidation states and can be oxidized or reduced. Hence, folate can exist in various forms namely N10-formyl THF (most oxidized form), N5, N10-methynyl THF, N5, N10-methylene THF, and N5-methyl THF (most reduced form). All these different forms carrying these single carbon groups are collectively known as the "one-carbon pool”.
Histidine and Tryptophan are other less common one-carbon sources that result in the formation of N5, N10-methynyl THF, and N10-formyl THF respectively. These forms are interconvertible, however, once a methyl group is formed, it is not readily oxidized back to N5, N10-methylene THF and hence N5-methyl THF tends to accumulate in the cells.
Folate is a B vitamin present in green leafy vegetables, fruits, and legumes in the diet as polyglutamate. The synthetic analog of folate is folic acid. It is also a synthetic oxidized dietary supplement that plays no direct biological role nor is it considered biologically active. It is absorbed in the jejunum and ileum after being reduced by folate reductase to N5-methyl THF which is the major form present in the blood and stored in the liver.
In the cells, folic acid is reduced to THF, a biologically active form, in a two-step process that requires two molecules of NADPH and the enzyme dihydrofolate reductase (DHFR).
A single carbon group is transferred from serine to THF by the enzyme serine hydroxymethyltransferase and reduced to N5, N10-methylene THF. Vitamin B6 is also required in this process. Glycine is produced as a result of this reaction, which can be readily reconverted to serine using the carbon from the one-carbon pool.
The carbon carried by N5, N10-methylene THF can then be transferred to deoxyuridine monophosphate (dUMP) to form deoxythymidine monophosphate (dTMP) by the enzyme thymidylate synthase that can be used to synthesize thymidine, at the same time recycling THF to DHF. Resulting DHF can be reduced once again to tetrahydrofolate and N5, N10-methylene THF again to continue this cycle. 5-fluorouracil is an anti-cancer drug that is used to inhibit the proliferation of cancer cells. It inhibits the synthesis of thymine and consequently DNA synthesis, by inhibiting the enzyme thymidylate synthase.
Alternately, N5, N10-methylene THF can be reduced to N5-methyl-THF, a storage form of folate, with the help of riboflavin (vitamin B2) and the enzyme methylenetetrahydrofolate reductase (MTHFR).
The methyl group from N5-methyl-THF is transferred to cobalamine (B12) to help regenerate methylcobalamin (B12) and THF. Methylcobalamine can then donate the methyl group to homocysteine and form methionine with the help of methionine synthase. Hence B12 deficiency results in increased levels of homocysteine in the blood. Additionally, methylmalonic acid levels also increase in B12 deficiency as the conversion of methylmalonyl CoA to Succinyl CoA is catalyzed by a vitamin B12 dependent enzyme, methylmalonyl CoA mutase. Folate deficiency does not have any effect on methylmalonic acid levels, but homocysteine levels increase as homocysteine cannot be converted to methionine due to deficiency of N5-methyl THF. B12 plays an important role in the recycling of N5-methyl THF (the storage form of folate) to THF. Deficiency of Vitamin B12 can impair this process and prevent the use of storage form of folate when needed, resulting in functional folate deficiency.
Serine and glycine cycle: Serine is a non-essential amino acid that can be obtained from supplements and in our diet. Glycine can form in our tissues from serine. Serine is the primary source of carbon in the conversion of tetrahydrofolate to N5, N10-methylene tetrahydrofolate. Glycine production also occurs in this reaction. The formation of serine requires a hydroxymethyl group from N5, N10-methylene tetrahydrofolate, and a glycine residue in the reverse reaction. 
Methionine is an important amino acid that is converted to S-adenosyl methionine (SAM) that participates in various methylation reactions and results in the formation of S-adenosyl homocysteine (SAH). SAM acts as a methyl donor, where it can donate single carbons to assist with the production of nucleic acids, proteins and neurotransmitters, and other methyltransferase reactions. Next the adenosyl group is removed from SAH to form homocysteine. A methyl group is donated by methylcobalamin (B12) to homocysteine resulting in the formation of methionine. This reaction also requires the enzyme methionine synthase. Resultant cobalamin (without the methyl group) can be regenerated back to methylcobalamin by accepting the methyl group from N5-methyl THF. This process allows the recycling of N5-methyl THF to THF for reuse.
Methotrexate and Folic acid deficiency: Methotrexate, a folic acid antagonist, is commonly used in cancer chemotherapy, inflammatory, and autoimmune conditions such as rheumatoid arthritis. As an irreversible competitive inhibitor of dihydrofolate reductase, it blocks the conversion of dihydrofolate (DHF) to tetrahydrofolate (THF) and therefore prevents cellular proliferation by preventing the synthesis of thymidine, a building block for nucleic acid synthesis and therefore ultimately inhibiting DNA and RNA synthesis. Therefore, all patients receiving methotrexate are given supplemental folic acid to prevent hematologic, gastrointestinal, and hepatic side effects. 
Megaloblastic Anemia: Deficiency of either folic acid or cobalamin (vitamin B12) can result in megaloblastic anemia, due to inhibition of DNA synthesis caused by decreased availability of purines and pyrimidines (Thymidine monophosphate). This results in enlarged red blood cells and accumulation of large, immature precursors (megaloblasts) of RBCs in the blood and bone marrow. Deficiency manifests in the form of fatigue, muscle weakness, tingling of extremities, and loss of joint position/coordination. 
Neural tube Defects: Folic acid deficiency is a known cause of neural tube defects such as spina bifida and anencephaly. During the first few weeks of fetal development, folic acid is a critical requirement for several folate-dependent processes. Therefore it is important that pregnant women and those who are planning to conceive, ensure adequate folic acid consumption.
Methylenetetrahydrofolate reductase (MTHFR) deficiency: Mutation in the MTHFR gene results in elevated plasma levels of homocysteine (hyperhomocysteinemia) as a defect in this enzyme prevents the conversion of homocysteine to methionine. MTHFR is the rate-limiting enzyme that catalyzes the formation of 5-methyltetrahydrofolate from N5, N10-methylene tetrahydrofolate, a cosubstrate for methylation of homocysteine to methionine. Genetic testing is available to identify this condition. It is important to diagnose hyperhomocysteinemia early since it is a risk factor for atherosclerosis, venous thrombosis, myocardial infarction, and other cardiovascular conditions. In addition, there are known polymorphisms associated with the MTHFR gene, that are associated with conditions such as colon cancer. 
Antibiotics: Trimethoprim and sulfamethoxazole are antibiotic drugs used in combination for the treatment of certain bacterial infections. Trimethoprim inhibits DHFR and prevents the synthesis of THF. Sulfamethoxazole is a structural analog of para-aminobenzoic acid that inhibits the bacterial synthesis of DHF.
|||Locasale JW, Serine, glycine and one-carbon units: cancer metabolism in full circle. Nature reviews. Cancer. 2013 Aug; [PubMed PMID: 23822983]|
|||Gonen N,Assaraf YG, Antifolates in cancer therapy: structure, activity and mechanisms of drug resistance. Drug resistance updates : reviews and commentaries in antimicrobial and anticancer chemotherapy. 2012 Aug [PubMed PMID: 22921318]|
|||Stover PJ, One-carbon metabolism-genome interactions in folate-associated pathologies. The Journal of nutrition. 2009 Dec; [PubMed PMID: 19812215]|
|||Carreras CW,Santi DV, The catalytic mechanism and structure of thymidylate synthase. Annual review of biochemistry. 1995; [PubMed PMID: 7574499]|
|||McBreairty LE,Bertolo RF, The dynamics of methionine supply and demand during early development. Applied physiology, nutrition, and metabolism = Physiologie appliquee, nutrition et metabolisme. 2016 Jun [PubMed PMID: 27177124]|
|||Hagner N,Joerger M, Cancer chemotherapy: targeting folic acid synthesis. Cancer management and research. 2010 Nov 19; [PubMed PMID: 21301589]|
|||Funk RS,van Haandel L,Becker ML,Leeder JS, Low-dose methotrexate results in the selective accumulation of aminoimidazole carboxamide ribotide in an erythroblastoid cell line. The Journal of pharmacology and experimental therapeutics. 2013 Oct [PubMed PMID: 23887097]|
|||Shea B,Swinden MV,Ghogomu ET,Ortiz Z,Katchamart W,Rader T,Bombardier C,Wells GA,Tugwell P, Folic acid and folinic acid for reducing side effects in patients receiving methotrexate for rheumatoid arthritis. The Journal of rheumatology. 2014 Jun [PubMed PMID: 24737913]|
|||Yadav MK,Manoli NM,Madhunapantula SV, Comparative Assessment of Vitamin-B12, Folic Acid and Homocysteine Levels in Relation to p53 Expression in Megaloblastic Anemia. PloS one. 2016; [PubMed PMID: 27780269]|
|||Aslinia F,Mazza JJ,Yale SH, Megaloblastic anemia and other causes of macrocytosis. Clinical medicine [PubMed PMID: 16988104]|
|||van der Put NM,Thomas CM,Eskes TK,Trijbels FJ,Steegers-Theunissen RP,Mariman EC,De Graaf-Hess A,Smeitink JA,Blom HJ, Altered folate and vitamin B12 metabolism in families with spina bifida offspring. QJM : monthly journal of the Association of Physicians. 1997 Aug [PubMed PMID: 9327028]|
|||Ebara S, Nutritional role of folate. Congenital anomalies. 2017 Sep; [PubMed PMID: 28603928]|
|||Guttormsen AB,Ueland PM,Nesthus I,Nygård O,Schneede J,Vollset SE,Refsum H, Determinants and vitamin responsiveness of intermediate hyperhomocysteinemia (> or = 40 micromol/liter). The Hordaland Homocysteine Study. The Journal of clinical investigation. 1996 Nov 1 [PubMed PMID: 8903338]|
|||Liew SC,Gupta ED, Methylenetetrahydrofolate reductase (MTHFR) C677T polymorphism: epidemiology, metabolism and the associated diseases. European journal of medical genetics. 2015 Jan [PubMed PMID: 25449138]|
|||[PubMed PMID: 6985448]|