Folate moiety

Fig. 6.22. Folate-FRET sensor structure and its application to measure disulfide bond reduction in endosomes. The molecule contains the folate moiety which is recognized by the folate receptor situated at the plasma membrane. This recognition leads to endocytosis and after some time to cleavage of the probe. 

The dihydrofolate reductase inhibitor, methotrexate (Fig. 8.47), was developed as an anticancer drug, whose inhibition of formation of folic acid coenzymes would block purine synthesis. In other words, it was designed to induce a folic acid deficiency. Notice in Figs. 8.50 and 8.51 that formation of dTMP, FGAR, and AICAR also causes the oxidation of tetra-hydrofolate to dihydrofolate. The latter must be reduced by dihydrofolate reductase to tet-rahydrofolate before active coenzyme can form again. Thus, not only does methotrexate inhibit the initial formation of the tetrahydro-folate moiety, it blocks regeneration of the coenzyme form. 

Coenzymes serve as recyclable shuttles—or group transfer reagents—that transport many substrates from their point of generation to their point of utilization. Association with the coenzyme also stabilizes substrates such as hydrogen atoms or hydride ions that are unstable in the aqueous environment of the cell. Other chemical moieties transported by coenzymes include methyl groups (folates), acyl groups (coenzyme A), and oligosaccharides (dolichol). 

A later paper65 reported a fluorescence maximum for leucovorin at 365 nm when excited at 314 nm in a pH 7 solution the concentration was 5 x 10-5 M. Variation between these data and other values was attributed to sample impurity, pH of solution, and quenching. The authors made an attempt to correlate structure and fluorescence of reduced folates. Similarity between tested compounds and jj-aminobenzoyl-glutamate lead them to conclude that this portion of the molecule is responsible for maxima at 360-425 nm when excited at 300-320 nm. They suggested that intensity differences may arise from various substitutions on the tetra-hydropteridine moiety. 

The oldest example of the use of nonclassical isosteres involves the replacement of the carboxamide in foUc acid by sulfonamide, to give the sulfanilamides. Diaminopyrimidines, as antimalarial agents, are also based on folate isosterism, in addition to the exploitation of auxiliary binding sites on dihydrofolate reductase. This concept of nonclassical isosteres or bioisosteres — that is, moieties that do not have the same nnmber of atoms or identical electron structure — is really the classical structure modification approach. 

The mode of action of sulfanilamides became known around 1947, when the structure and biosynthesis of folic acid were elucidated. This compound is built by bacteria from the heterocyclic pteroyl moiety, p-aminobenzoate, and glutamate. p-Aminobenzene-sulfonamide (9.89, sulfanilamide) is a competitive inhibitor of the synthase enzyme, acting as an antimetabolite of p-aminobenzoate. Occasionally, the sulfanilamide can even be incorporated into the modified folate, resulting in an inactive compound and thus an inactive enzyme. This theory, proposed by Woods and Fildes in 1940, became the first molecular explanation of drug action. 

Figure 14.2 Chemical structure of folates. Folate molecules consist of pteridine, para-aminobenzoate (pABA), and glutamate moieties. Plants usually contain polyglutamylated forms of folates that are made by the addition of up to about six glutamate residues (which form the y-glutamate tail) attached to the first glutamate, each linked by amide bonds to the preceding molecule of glutamate through the y-carboxyl of the latter. Cl units at various levels of oxidation can be attached to NS and/or N1 0, as indicated by Ri and R2.

The pteroic acid moiety of tetrahydrofolate consists of a reduced pteridine ring and p-aminobenzoic acid. Folate from the diet is absorbed by the intestinal mucosa and in two enzymatic 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 readily do so. 

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