Folic acid binding activity

Competitive inhibitors bind to specific groups in the enzyme active site to form an enzyme-inhibitor complex. The inhibitor and substrate compete for the same site, so that the substrate is prevented from binding. This is usually because the substrate and inhibitor share considerable stmctural similarity. Catalysis is diminished because a lower proportion of molecules have a bound substrate. Inhibition can be relieved by increasing the concentration of substrate. Some simple examples are shown below. Thus, sulfanilamide is an inhibitor of the enzyme that incorporates j9-aminobenzoic acid into folic acid, and has antibacterial properties by restricting folic acid biosynthesis in the bacterium (see Box 11.13). Some phenylethylamine derivatives, e.g. phenelzine, provide useful antidepressant drags by inhibiting the enzyme monoamine oxidase. The cA-isomer maleic acid is a powerful inhibitor of the enzyme that utilizes the trans-isomer fumaric acid in the Krebs cycle. 

Folic Acid Antagonists. Folic acid antagonists block the biosynthesis of purine nucleotides. Methotrexate (7.76) is the prototypic fohc acid antagonist and functions by binding to the active catalytic site of dihydrofolate reductase, thereby interfering with the synthesis of the reduced form that accepts one-carbon units lack of this cofactor blocks the synthesis of purine nucleotides. As well as being used in the treatment of cancer, methotrexate has been used in the management of rheumatoid arthritis, psoriasis, and even asthma. 

Methotrexate (Amethopterin) is a folic acid antagonist that binds to dihydrofolate reductase, thus interfering with the synthesis of the active cofactor tetrahydrofolic acid, which is necessary for the synthesis of thymidylate, purine nucleotides, and the amino acids serine and methionine. 

Animals are unable to synthesize folic acid (6.62) and must consume adequate quantities in their diets. Plants and bacteria, however, are able to make folic acid. The first step of this synthesis is catalyzed by dihydropteroate synthetase and reacts dihydroptero-ate diphosphate (6.69) and para-aminobenzoic acid (PABA, 6.70) (Figure 6.25). Because this pathway is not found in humans, inhibition of the reaction is a method to ultimately stop TMP synthesis in an invading bacterium while not impacting the infected host. The sulfonamides, often called sulfa drugs, are a class of antibiotic that exploits the folic acid pathway and inhibits dihydropteroate synthetase. Sulfa drugs bind in the same fashion as PABA and act as competitive inhibitors. The active form of the first sulfa drug is sulfanilamide (6.71). Sulfamethoxazole (6.72) is a sulfa drug that is widely prescribed today.26... 

Some of the structural properties which have been ascribed to DA receptors appear to deserve attention for their heuristic value, but painfully few should engender much confidence in their reality. A sobering lesson is available from analysis of complexes of dihydrofolate reductase (], 8). Methotrexate is a very close analog of folic acid and is a potent inhibitor of the enzyme, but it is now almost certain that these ligands bind in the enzyme active site in aspects differing by a rota-... 

For a displacement interaction to become clinically important, a second mechanism usually operates sodium valproate can cause phenytoin toxicity because it both displaces phenytoin from its binding site on plasma albumin and inhibits its metabolism. Similarly aspirin and probenecid (and possibly other nonsteroidal anti-inflammatory drugs) displace the folic acid antagonist methotrexate from its protein-binding site and reduce its rate of active secretion by the renal tubules the result is serious methotrexate toxicity. 

Sulfonamides act as inhibitors by mimicking p-aminobenzoic acid (PABA) (Fig. 10.14)—one of the normal constituents of folic acid. The sulfonamide molecule is similar enough in structure to PABA that the enzyme is fooled into accepting it into its active site (Fig. 10.15). Once it is bound, the sulfonamide prevents PABA from binding. As a result, folic acid is no longer synthesized. Since folic acid is essential to cell growth, the cell will stop dividing. 

Receptor-binding activity of the protein-folate conjugate can be determined by competitive displacement of a known quantity of [3H]-labeled folic acid (available from NEN Life Science Products, Boston, MA) from receptor-expressing cells. Alternatively, if the protein can be labeled with a fluorescent probe (e.g., fluorescein isothiocyanate), its uptake by folate receptor-bearing cells can be monitored by fluorescence microscopy (8). 

If the correct substrate (PABA) is bound by the enzyme, the reaction occurs, and the bacterium lives. However, if the sulfa drug is present in excess over PABA, it binds more frequently to the active site of the enz)une. No folic acid will be produced, and the bacterial cell will die. 

Vitamin Bj2 may be found in liver, kidney, fish, and fortified milk and helps convert folic acid into its active form. Vitamin Bj2 is essential to synthesize DNA and promotes cellular division and is required for hematopoiesis (development of red blood cells in bone marrow) and to maintain the integrity of the nervous system. Vitamin Bj2 is absorbed in the intestine with the aid of an intrinsic factor produced by gastric parietal cells. Once absorbed, vitamin Bj2 binds to... 

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