N5,N10-methylene tetrahydrofolate

Tetrahydrofolate accepts methyl groups, usually from serine. The product, N5,N10-methylene-tetrahydrofolate, is the central compound in 1-carbon metabolism. Tetrahydrofolate can also accept a methyl group from the complete breakdown of glycine. In Figure 5-2, only the N5, C6, and N10 atoms of the pteroic acid are shown for clarity. 

The donation of the methyl group from N5,N10-methylene tetrahydrofolate leads to the oxidation of the cofactor to dihydrofolate. This points to the importance of dihydrofolate reductase (DHFR) in the functioning of thymidylate synthase. Thus, synthesis of TMP requires a supply of both methyl groups—for example, from serine— and reducing equivalents. 

Site of action 5-FU per se is devoid of antineoplastic activity and must be converted to the corresponding deoxynucleotide (5-FdUMP, Figure 38.9), which competes with deoxyuridine monophosphate (dUMP) for thymidylate synthetase. 5-FdUMP acts as a pseudosubstrate and is entrapped with the enzyme and its N5,N10-methylene tetrahydrofolic acid coenzyme in a ternary complex that cannot proceed to products. DNA synthesis decreases due to lack of thymidine, leading to imbalanced cell growth and cell death. [Note Leucovorin is given with 5-FU because the reduced folate coenzyme is required in the thymidylate synthetase reaction. Lack of sufficient coenzyme reduces the effectiveness of the antipyrimidine.] 5-FU is also incorporated into RNA and low levels have been detected in DNA. 

The answer is c. (Murray, pp 48-73. Scriver, pp 4571-4636. Sack, pp 3-17. Wilson, pp 287-317.) Since rapidly multiplying cancer cells are dependent upon the synthesis of deoxythymidilate (dTMP) from deoxy-uridylate (dUMP), a prime target in cancer therapy has been inhibition of dTMP synthesis. The anticancer drug fluorouracil is converted in vivo to fluorodeoxyuridylate (FdUMP), which is an analogue of dlJMP FdUMP irreversibly forms a covalent complex with the enzyme thymidylate synthase and its substrate N5,N10-methylene-tetrahydrofolate. This is a case of suicide inhibition, where an enzyme actually participates in the change of a substrate into a covalently linked inhibitor that irreversibly inhibits its catalytic activity. 

Histidine can be degraded to form ammonia and formiminoglutamate. The latter, with tetrahydrofolate, can form ammonia, N5, N10 methylene tetrahydrofolate and glutamate. 

All the pyrimidine nucleotides can proceed to the deoxy compounds as diphosphates, similar to the purines. However, uridine does not occur in DNA therefore, it must be converted to a thymine derivative. This occurs by the following basic pathway CDP is converted to dCDP. This is converted to dCMP, which can produce dUMP. The dUMP is then methylated in the number 5 position, via N5, N10-methylene tetrahydrofolate. This produces deoxythymidine monophosphate (dTMP) and dihydrofolate. The dTMP then can be converted to dTDP and dTTP, which can be incorporated into DNA (Fig. 20.9). 

The effects of cobalamin deficiency are most pronounced in rapidy dividing cells, such as the erythropoietic tissue of bone marrow and the mucosal cells of the intestine. Such tissues need both Die N5-N10-methylene and N10-formyl forms of tetrahydrofolate for Ihe synthesis of nucleotides required for DNA replication (see pp. 291, 301). However, in vitamin B12 deficiency, the N5-methyl form of tetrahydrofolate is not efficiently used. Because the methylated fonn cannot be converted directly to other forms of tetrahydrofolate, tie Ns-methyl form accumulates, whereas the levels of the other forms decrease. Thus, cobalamin deficiency is hypothesized to lead to a deficiency of the tetrahydrofolate forms needed in purine and thymine synthesis, resulting in the symptoms of megaloblastic anemia. 

Hong, B., and Kohen, A., 2005. Microscale synthesis of isotopically labeled (6R)-N5,N10-methylene-5,6,7,8-tetrahydrofolate. Journal of Labelled Compounds Radiopharmaceuticals. 48 759-769. 

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