Folate (folic acid → DHF → THF) is reduced via DHFR — the methotrexate target. THF accepts one-carbon units (from serine, glycine, formate) at the N5/N10 positions to form: 5-methyl-THF (donates methyl to homocysteine → methionine), 5,10-methylene-THF (donates methyl to dUMP → dTMP via thymidylate synthase — the 5-FU target), 10-formyl-THF (donates formyl to purine de novo synthesis). Methotrexate inhibits DHFR + thymidylate synthase + AICAR transformylase — explaining its activity in chemotherapy (kills proliferating cells), rheumatology (immunosuppressant), and obstetrics (ectopic pregnancy + medical abortion). Leucovorin (5-formyl-THF) bypasses DHFR for rescue.
Organ Systems
immune-hematologic
reproductive
Pathway Steps
folic-acid → dihydrofolate — via folate reductase. Supplemental folic acid (the synthetic, fully oxidized form) must be reduced before use; this first reduction is slow, so high doses can leave unmetabolized folic acid in circulation — part of the rationale for supplementing reduced 5-MTHF instead.
dihydrofolate → tetrahydrofolate — via DHFR — METHOTREXATE TARGET. Dihydrofolate reductase regenerates tetrahydrofolate, the active one-carbon carrier, and is the target of methotrexate, trimethoprim (bacterial-selective) and pyrimethamine (antiparasitic) — species differences in DHFR give these drugs their selectivity.
tetrahydrofolate → 5-methyl-tetrahydrofolate — via methylene-THF reductase (MTHFR) — common C677T polymorphism. MTHFR commits one-carbon units to remethylating homocysteine to methionine (via B12-dependent methionine synthase). The common C677T variant lowers activity, modestly raising homocysteine; 5-MTHF is the form that crosses into the CNS and feeds the methionine cycle.
tetrahydrofolate → 5-10-methylene-tetrahydrofolate — via serine hydroxymethyltransferase (SHMT) — substrate for thymidylate synthase. Serine hydroxymethyltransferase moves a one-carbon unit from serine (a major one-carbon source) onto THF, giving the 5,10-methylene-THF used by thymidylate synthase for dTMP — linking folate status directly to DNA synthesis and to the antifolate mechanism.
pyridoxine (cofactor) — SHMT (serine ↔ glycine + 5,10-methylene-THF). B6 (pyridoxine) converts to PLP, the cofactor for serine hydroxymethyltransferase — the enzyme that generates one-carbon units onto THF from serine.