Dihydrofolate Reductase

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Dihydrofolate reductase uses NADPH to reduce dihydrofolate to produce tetrahydrofolate oxidoreductase. Dihydrofolate reductase that exists in microorganisms and liver has been studied in detail. The molecular weight is about 20,000. It also seems to have the effect of catalyzing the reduction of folic acid to dihydrofolate, so folic acid can be turned into an active form of tetrahydrofolate for the enzyme coenzyme.

Figure 1. Protein structure of dihydrofolate reductase.

Introductions

Dihydrofolate reductase (DHFR) catalyses the NADPH-dependent reduction of dihydrofolate to tetrahydrofolate, an essential step in de novo synthesis both of glycine and of purines and deoxythymidine phosphate (the precursors of DNA synthesis), and important also in the conversion of deoxyuridine monophosphate to deoxythymidine monophosphate. Although DHFR is found ubiquitously in prokaryotes and eukaryotes, and is found in all dividing cells, maintaining levels of fully reduced folate coenzymes, the catabolic steps are still not well understood. Bacterial species possesses distinct DHFR enzymes (based on their pattern of binding diaminoheterocyclic molecules), but mammalian DHFRs are highly similar. The active site is situated in the N-terminal half of the sequence, which includes a conserved Pro-Trp dipeptide; the tryptophan has been shown to be involved in the binding of substrate by the enzyme. Its central role in DNA precursor synthesis, coupled with its inhibition by antagonists such as trimethoprim and methotrexate, which are used as anti-bacterial or anti-cancer agents, has made DHFR a target of anticancer chemotherapy. However, resistance has developed against some drugs, as a result of changes in DHFR itself

Structure

The central eight-stranded beta sheet constitutes the main feature of DHFR polypeptide backbone folding. Seven of these chains are parallel and the eighth is antiparallel. Four alpha helices connect the continuous beta chain. Residues 9-24 are called "Met20" or "loop 1", and together with other loops, are part of the main subdomain surrounding the active site. The active site is located in the N-terminal half of the sequence and includes a conservative Pro-Trp dipeptide; tryptophan has been shown to be involved in the substrate binding of the enzyme.

Clinical significance

Dihydrofolate reductase deficiency is related to megaloblastic anemia and can be treated by reducing the form of folic acid. Since tetrahydrofolate, the product of this reaction, is the active form of human folic acid, inhibition of DHFR can lead to functional folic acid deficiency. Because DHFR plays a key role in the synthesis of DNA precursors, it is an attractive inhibitory drug. Trimethoprim is an antibiotic that inhibits bacterial DHFR, while methotrexate (a chemotherapy drug) can inhibit mammalian DHFR. However, due to mutations in DHFR itself, resistance to certain drugs has developed. Mutations in DHFR lead to rare autosomal recessive congenital folate metabolism errors, which can lead to megaloblastic anemia, pancytopenia and severe folate deficiency. Can be corrected and supplemented by leucovorin.

Functions

Dihydrofolate reductase converts dihydrofolate into tetrahydrofolate, which is a methyl shuttle required for the de novo synthesis of purines, thymidylate and certain amino acids. Although the functional dihydrofolate reductase gene has been mapped to chromosome 5, multiple pseudogenes or dihydrofolate reductase-like genes without intron processing have been identified on different chromosomes. Found in all organisms, DHFR plays a vital role in regulating the amount of tetrahydrofolate in cells. Tetrahydrofolate and its derivatives are essential for the synthesis of purine and thymidylic acid, and are important for cell proliferation and cell growth. DHFR plays a central role in the synthesis of nucleic acid precursors, and it has been shown that mutant cells completely lacking DHFR require glycine, amino acids and thymidine to grow. DHFR has also been shown to be an enzyme involved in the rescue of tetrahydrobiopterin from dihydrobiopterin.