Dihydropteroate

Subsequent knowledge of the stmcture, function, and biosynthesis of the foHc acid coenzyme gradually allowed a picture to be formed regarding the step in this pathway that is inhibited by sulfonamides. The biosynthetic scheme for foHc acid is shown in Figure 1. Sulfonamides compete in the step where condensation of PABA with pteridine pyrophosphate takes place to form dihydropteroate (32). The amino acids, purines, and pyrimidines that are able to replace or spare PABA are those with a formation that requkes one-carbon transfer catalyzed by foHc acid coenzymes (5). 

Development of Resistance. One of the principal disadvantages of sulfonamide therapy is the emergence of dmg-resistant strains of bacteria. Resistance develops by several mechanisms overproduction of PABA (38) altered permeabiUty of the organisms to sulfonamides (39) and reduced affinity of dihydropteroate synthetase for sulfonamides while the affinity for PABA is retained (40). Sulfonamides also show cross-resistance to other sulfonamides but not to other antibacterials. In plasmodia, resistance may occur by means of a bypass mechanism in which the organisms can use preformed foHc acid (41). 

In E. coli GTP cyclohydrolase catalyzes the conversion of GTP (33) into 7,8-dihydroneoptetin triphosphate (34) via a three-step sequence. Hydrolysis of the triphosphate group of (34) is achieved by a nonspecific pyrophosphatase to afford dihydroneopterin (35) (65). The free alcohol (36) is obtained by the removal of residual phosphate by an unknown phosphomonoesterase. The dihydroneoptetin undergoes a retro-aldol reaction with the elimination of a hydroxy acetaldehyde moiety. Addition of a pyrophosphate group affords hydroxymethyl-7,8-dihydroptetin pyrophosphate (37). Dihydropteroate synthase catalyzes the condensation of hydroxymethyl-7,8-dihydropteroate pyrophosphate with PABA to furnish 7,8-dihydropteroate (38). Finally, L-glutamic acid is condensed with 7,8-dihydropteroate in the presence of dihydrofolate synthetase. 

Dihydropteroic acid (85) is an intermediate to the formation of the folic acid necessary for intermediary metabolism in both bacteria and man. In bacteria this intermediate is produced by enzymatic condensation of the pteridine, 86, with para-amino-benzoic acid (87). It has been shown convincingly that sulfanilamide and its various derivatives act as a false substrate in place of the enzymatic reaction that is, the sulfonamide blocks the reaction by occupying the site intended for the benzoic acid. The lack of folic acid then results in the death of the microorganism. Mammals, on the other hand, cannot synthesize folic acid instead, this compound must be ingested preformed in the form of a vitamin. Inhibition of the reaction to form folic acid Ls thus without effect on these higher organisms. 

Overproduction of the chromosomal genes for the dihydrofolate reductase (DHFR) and the dihydroptero-ate synthase (DHPS) leads to a decreased susceptibility to trimethoprim and sulfamethoxazol, respectively. This is thought to be the effect of titrating out the antibiotics. However, clinically significant resistance is always associated with amino acid changes within the target enzymes leading to a decreased affinity of the antibiotics. 

There is another fundamental difference between folate utilization in microbial and mammalian cells. Bacteria and protozoa are unable to take up exogenous folate and must synthesize it themselves. This is carried out in a series of reactions involving first the synthesis of dihydropteroic acid from one molecule each of pteridine and p-aminobenzoic acid (PABA). Glutamic acid is then added to form DHF which is reduced by DHFR to THF. Mammalian cells do not make their own DHF, instead they take it up firm dietary nutrients and convert it to THF using DHFR. 

Sulphonamides are structural analogues of PABA. They competitively inhibit the incorporation of PABA into dihydropteroic acid and there is some evidence for their incorporation into false folate analogues which inhibit subsequent metabolism. The presence of excess PABA will reverse the inhibitory action of sulphonamides, as will thymine, adenine, guanine and methionine. However, these nutrients are not normally available at the site of infections for which the sulphonamides are used. 

Two mechanisms of chromosomal resistance have been identified. A mutation of dihydropteroate synthetase (DHPS) in Strep, pneumoniae produces an altered enzyme with reduced affinity for sulphonamides. Hyperproduction of p-aminobenzoic acid (PABA) overcomes the block imposed by inhibition ofDHPS. The specific cause of PABA hyperproduction is unknown, though chromosomal mutation is the probable cause. 

Mouillon, J.M. et al., Folate synthesis in higher-plant mithocondria coupling between the dihydropterin pyrophosphokinase and the dihydropteroate synthase activities, Biochem. J., 363, 313, 2002. 

The macrolide erythromycin inhibits protein synthesis and resistance is induced by N -dimethyl-ation of adenine within the 23S rRNA, which results in reduced affinity of ribosomes for antibiotics related to erythromcin (Skinner et al. 1983). Sulfonamides function by binding tightly to chromosomal dihydropteroate synthetase and resistance to sulfonamides is developed in the resistance plasmid through a form of the enzyme that is resistant to the effect of sulfonamides. 

The answer is c. (Hardman, pp 1058-1059. Katzung, pp 793-795.) Trimethoprim inhibits dihydro folic acid reductase. Sulfamethoxazole inhibits p-aminobenzoic acid (PABA) from being incorporated into folic acid by competitive inhibition of dihydropteroate synthase. Either action inhibits the synthesis of tetrahydrofolic acid. 

Dihydrojasmone, 14 598 Dihydrolinalool, 24 502 Dihydromyrcene, 24 487, 488 Dihydromyrcenol, 24 487, 488, 495 Dihydroperoxides geminal, 18 455-456 thermal decomposition of, 18 455 Di(hydroperoxyalkyl) peroxides, 18 459 Dihydropteroate synthase (DHPS),... 

In contrast to humans, bacteria have the biochemical ability to synthesize folic acid from simpler molecules. Here we have a clear biochemical difference between human beings and infectious organisms that we can exploit to our benefit. The reaction catalyzed by an enzyme known as dihydropteroate synthetase, in which a complex heterocycle is linked to p-aminobenzoic acid, is key. Now recognize the structural similarity between sulfanilamide, or other sulfonamides, and p-aminobenzoic acid ... 

Given this structural similarity, it should not be surprising to learn that sulfanilamide competes with p-aminobenzoic acid for a binding site on the surface of dihydropteroate synthetase. Put another way, sulfanilamide binds to the enzyme where p-aminobenzoic acid should bind but no reaction occurs. The consequence is that a step in folic acid biosynthesis is disrupted and the bacterial cell is deprived of adequate folic acid. Nucleic acid synthesis, among other things, is disrupted, leading to a cessation of cell growth and division. The human immune system can mop up what remains. No similar consequences befall the human host since it cannot make folic acid in the first place and must get an adequate supply of this vitamin in the diet. 

Genes encoding phosphotransferases confer resistance to streptomycin Genes encoding a drug-resistant dihydropteroate synthase enzyme required for folate biosynthesis confer resistance to sulfonamide Tetracycline... 

Mammals must obtain their tetrahydrofolate requirements from their diet, but microorganisms are able to synthesize this material. This offers scope for selective action and led to the use of sulfanilamide and other antibacterial sulfa drugs, compounds that competitively inhibit the biosynthetic enzyme (dihydropteroate synthase) that incorporates p-aminobenzoic acid into the structure (see Box 7.23). 

This enzyme [EC 6.3.2.12] catalyzes the reaction of ATP with dihydropteroate and glutamate to generate ADP, orthophosphate, and dihydrofolate. 

This enzyme [EC 2.5.1.15], also known as dihydro-pteroate pyrophosphorylase, catalyzes the reaction of 2-amino-4-hydroxy-6-hydroxymethyl-7,8-dihydropteri-dine diphosphate with 4-ammobenzoate to produce pyrophosphate and dihydropteroate. 

DIHYDROOROTATE DEHYDROGENASE DIHYDROOROTATE OXIDASE DIHYDROPTERIDINE REDUCTASE DIHYDROPTEROATE SYNTHASE DIHYDROPYRIMIDINASE DIHYDROURACIL DEHYDROGENASE Dihydroxyacetone kinase,... 

Dienestrol, 102, 103 Diethyl carbamazine, 278 Diethylstilbestrol, 101 Diethylthiambutene, 106 Dihexyrevine, 36 Dihydralazine, 353 Dihydrocodeine, 288 Dihydropteroic acid, in bacterial metabolism, 121 Diketene, addition to imines,... 

Thus sulfonamides are bacteriostatic drugs that inhibit bacterial growth by interfering with the microbial synthesis of folic acid. More specifically, sulfonamides block the biosynthetic pathway of folic acid synthesis, thus competitively inhibiting the transformation of p-aminobenzoic acid to folic acid (mediated by the enzyme dihydropteroate synthetase), which allows them to be considered as antimetabolites. 

About 10-25%, i.e. 50-200 pg, of the daily dietary intake of folic acid in yeasts, liver, and green vegetables is absorbed via active and passive transport in the proximal jejunum. As humans do not have dihydropteroate synthetase, which synthesizes folic acid in bacteria, we require folic acid in the diet. Only small amounts of folate can be stored in the body and dietary deficiency for only a few days can result in symptomatic folate deficiency. 

Both the sulfonamides and trimethoprim interfere with bacterial folate metabolism. For purine synthesis tetrahydrofolate is required. It is also a cofactor for the methylation of various amino acids. The formation of dihydrofolate from para-aminobenzoic acid (PABA) is catalyzed by dihydropteroate synthetase. Dihydrofolate is further reduced to tetrahydrofolate by dihydrofolate reductase. Micro organisms require extracellular PABA to form folic acid. Sulfonamides are analogues of PABA. They can enter into the synthesis of folic acid and take the place of PABA. They then competitively inhibit dihydrofolate synthetase resulting in an accumulation of PABA and deficient tetrahydrofolate formation. On the other hand trimethoprim inhibits dihydrofolate... 

Both sulfonamides and trimethoprim (not a sulfonamide) sequentially interfere with folic acid synthesis by bacteria. Folic acid functions as a coenzyme in the transfer of one-carbon units required for the synthesis of thymidine, purines, and some amino acids and consists of three components a pteridine moiety, PABA, and glutamate (Fig. 44.1). The sulfonamides, as structural analogues, competitively block PABA incorporation sulfonamides inhibit the enzyme dihydropteroate synthase, which is necessary for PABA to be incorporated into dihydropteroic acid, an intermediate compound in the formation of folinic acid. Since the sulfonamides reversibly block the synthesis of folic acid, they are bacteriostatic drugs. Humans cannot synthesize folic acid and must acquire it in the diet thus, the sulfonamides selectively inhibit microbial growth. 

Resistance to the sulfonamides can be the result of decreased bacterial permeability to the drug, increased production of PABA, or production of an altered dihydropteroate synthetase that exhibits low affinity for sulfonamides. The latter mechanism of resistance is plasmid mediated. Active efflux of the sulfonamides has also been reported to play a role in resistance. The inhibitory effect of the sulfonamides also can be reversed by the presence of pus, tissue fluids, and drugs that contain releasable PABA. 

E) Structural changes in dihydropteroate synthase and overproduction of PABA. 

B. Humans cannot synthesize folic acid (A) diet is their main source. Sulfonamides selectively inhibit microbially synthesized folic acid. Incorporation (B) of PABA into microbial folic acid is competitively inhibited by sulfonamides. The TMP-SMX combination is synergistic because it acts at different steps in microbial folic acid synthesis. All sulfonamides are bacteriostatic. Inhibition of the transpeptidation reaction (C) involved in the synthesis of the bacterial cell wall is the basic mechanism of action of (3-lac-tam antibiotics Changes in DNA gyrases (D) and active efflux transport system are mechanisms for resistance to quinolones. Structural changes (E) in dihydropteroate synthetase and overproduction of PABA are mechanisms of resistance to the sulfonamides. 

Whereas the sulfonamides and sulfones inhibit the initial step whereby PABA and the pteridine moiety combine to form dihydropteroic acid (see Chapter 44), pyrimethamine and trimethoprim inhibit the conversion of dihydrofolic acid to tetrahydrofoUc acid, a reaction... 

Inhibit folic acid synthesis by competitive inhibition of dihydropteroate synthase. 

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