NADPH folate reduction

Folic acid and its polyglutamyl derivatives can be reduced to the THF coenzymes in two stages the first step is a slow reduction with NADPH to 7,8-dihydro-folate (step a, Fig. 15-18). The same enzyme that catalyzes this reaction rapidly reduces the dihydrofolates... 

Methylene-Tetrahydrofolate Reductase The reduction of methylene-tetrahydrofolate to methyl-tetrahydrofolate, shown in Figure 10.7, is catalyzed hy methylene-tetrahydrofolate reductase, a flavin adenine dinucleotide-dependent enzyme during the reaction, the pteridine ring of the substrate is oxidized to dihydrofolate, then reduced to tetrahydrofolate by the flavin, which is reduced by nicotinamide adenine dinucleotide phosphate (NADPH Matthews and Daubner, 1982). The reaction is irreversible under physiological conditions, and methyl-tetrahydrofolate - which is the main form of folate taken up into tissues (Section 10.2.2) - can only be utilized after demethylation catalyzed by methionine synthetase (Section 10.3.4). 

The metabolism of folic acid involves reduction of the pterin ting to different forms of tetrahydrofolylglutamate. The reduction is catalyzed by dihydtofolate reductase and NADPH functions as a hydrogen donor. The metabolic roles of the folate coenzymes are to serve as acceptors or donors of one-carbon units in a variety of reactions. These one-carbon units exist in different oxidation states and include methanol, formaldehyde, and formate. The resulting tetrahydrofolylglutamate is an enzyme cofactor in amino acid metabolism and in the biosynthesis of purine and pyrimidines (10,96). The one-carbon unit is attached at either the N-5 or N-10 position. The activated one-carbon unit of 5,10-methylene-H folate (5) is a substrate of T-synthase, an important enzyme of growing cells. 5-10-Methylene-H folate (5) is reduced to 5-methyl-H,j folate (4) and is used in methionine biosynthesis. Alternatively, it can be oxidized to 10-formyl-H folate (7) for use in the purine biosynthetic pathway. 

NADPH and dihydrofolate reductase convert folate to dihydrofolate (FH2), which may undergo a second reduction by the same enzyme to form FH4. 

In the tissues, tetrahydrofolate is converted to polyg-lutamyl forms by an ATP-dependent synthetase. In the liver, the major form is pteroyl pentaglutamate. Reduced polyglutamyl forms, each substituted with one of several one-carbon moieties, are the preferred coenzymes of folate-dependent enzymes. Reduction of folate (F) to tetrahydrofolate (FH4) occurs in two steps F is reduced to 7,8-dihydrofolate (FH2), and FH2 is reduced to 5,6,7,8-tetrahydrofolate (FH4). Both of these reactions are catalyzed by a single NADPH-linked enzyme, dihydrofolate reductase (Figure 27-2). 

The stereochemistry of the reduction of dihydrofolate to H4-folate by the enzyme requires transfer of hydrogen at the re face of dihydrofolate, 3, to generate the correct stereochemistry at C-6. This transfer involves the 4-pro-R hydrogen of NADPH for the reductases from chicken liver, L1210 cells, S. faecalis and E. coli [35-38]. The reductase from L. casei will also reduce folic acid, albeit at a slower... 

The pteroic acid moiety of tetrahydrofolate consists of a reduced pteridine ring and j)-aminobenzoic acid. Folic acid from the diet is absorbed by the intestinal mucosa, and in two enzymic steps is reduced to tetrahydrofolate which is the active form of the coenzyme. Mammals cannot synthesize folate this normally does not present a problem because microorganisms of the intestinal tract do so in sufficient quantities. The two steps in the reduction of folic acid to tetrahydrofolate are catalyzed by dihydrofolate reductase. Both of these reactions require NADPH as a somce of electrons. 

The enzyme dihydrofolate reductase (DHFR) catalyzes the nicotinamide adenine dinucleotide phosphate (NADPH) reduction of folate to dihydrofolate and tetrahydrofolate, the class of cofactors used in the biosynthesis of thymidylate and hence DNA. Inhibition of DHFR prevents cell growth and kills cells, so... 

Thus, the formation of thymidylate involves the formation of a carbon-carbon bond and a reduction 5,10-methyIene H4-folate is both donor of the one-carbon unit and reductant. It is apparent that a catalytic amount of H4-folate will serve in the cycle. This is consistent with the observation that in S. faecalis extracts the synthesis of thymidylate in the presence of excess H4-folate was unaffected by aminopterin, a potent inhibitor of tetrahydrofolate dehydrogenase when this reaction was conducted with catalytic amounts of H4-folate and excess NADPH, thymidylate synthesis was blocked by aminopterin (5). 

A rare case of enzyme catalyzing imine reduction reaction (see also Section 13.4.4) is the stereoselective reduction of dihydrofolic acid to (6S)-tetrahydrofolic acid by dihydrofolate reductase (DHFR) at the expense of NADPH. This biocatalytic step was employed in the synthesis of (S)-leucovorin [(6S)-5-formyl-5,6,7,8-tetrahydro-folate], a drug used in cancer chemotherapy. DHFR produced by E. coli was combined with a GDH/glucose cofactor recycling system and yielded (6S)-tetrahy-drofolic acid, which upon formylation furnished L-leucovorin with >99.5% de (Figure 13.31) [37-39]. 

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