Inhibition feedback

As outlined in sections II. 2-4, several enzymes are subject to negative feedback control during sialic acid formation. These are listed in Fig. 9. The central intermediates are UDP-GlcNAc and CMP-Neu5Ac. As has been described for other monosaccharide activation pathways, the negative feedback influences the first enzymic steps in the pathway (ScHACHTERand Roden 1973, Schachter 1978). 

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The production of elfamycins is described in the references cited in Table 1. Fermentation yield improvements with aurodox (1, R = CH ) proved difficult because of feedback inhibition (48). Aurodox-resistant strains (49), however, responded positively to conventional mutagenic methods leading to yield increases from 0.4 to 2.5 g/L (50). Scale-up of efrotomycin (7, R = CH ) fermentations were found to be particularly sensitive to small changes in sterilization conditions of the oil-containing medium used (51). 

Catecholamine biosynthesis begins with the uptake of the amino acid tyrosine into the sympathetic neuronal cytoplasm, and conversion to DOPA by tyrosine hydroxylase. This enzyme is highly localized to the adrenal medulla, sympathetic nerves, and central adrenergic and dopaminergic nerves. Tyrosine hydroxylase activity is subject to feedback inhibition by its products DOPA, NE, and DA, and is the rate-limiting step in catecholamine synthesis the enzyme can be blocked by the competitive inhibitor a-methyl-/)-tyrosine (31). 

Fermentation can be combined with other operations. For example, feedback inhibition of enzymatic hydrolysis of cellulose can be relieved by removal of the product glucose by fermentation as it forms. This is teni ed. simultaneou.s-saccharification-fermentation (SSF). 

In this scheme, F symbolizes an essential metabolite, such as an amino acid or a nucleotide. In such systems, F, the essential end product, inhibits enzyme 1, xAie first step in the pathway. Therefore, when sufficient F is synthesized, it blocks further synthesis of itself. This phenomenon is called feedback inhibition or feedback regulation. 

The small overproduction of amino adds by wild type strains in culture media is the result of regulatory mechanisms in the biosynthetic pathway. These regulatory mechanisms are feedback inhibition and repression. 

In die metabolic pathway to an amino add several steps are involved. Each step is die result of an enzymatic activity. The key enzymatic activity (usually die first enzyme in the synthesis) is regulated by one of its products (usually die end product, eg die amino add). If die concentration of die amino add is too high die enzymatic activity is decreased by interaction of die inhibitor with the regulatory site of die enzyme (allosteric enzyme). This phenomenon is called feedback inhibition. 

These mutants lack feedback inhibition and are used for the production of many amino adds. 

Selection of these regulatory mutants is often done by using toxic analogues of amino adds for example p-fluoro-DL-phenylalanine is an analogue of phenylalanine. Mutants that have no feedback inhibition or repression to the amino add are also resistant to the analogue amino add. They are therefore selected for and can be used to overproduce the amino add. Some amino add analogues function as false co-repressors, false feedback inhibitors or inhibit the incorporation of foe amino acid into foe protein. 

A possible explanation for the superiority of the amino donor, L-aspartic add, has come from studies carried out on mutants of E. coli, in which only one of the three transaminases that are found in E. coli are present. It is believed that a branched chain transaminase, an aromatic amino add transaminase and an aspartate phenylalanine aspartase can be present in E. coli. The reaction of each of these mutants with different amino donors gave results which indicated that branched chain transminase and aromatic amino add transminase containing mutants were not able to proceed to high levels of conversion of phenylpyruvic add to L-phenylalanine. However, aspartate phenylalanine transaminase containing mutants were able to yield 98% conversion on 100 mmol l 1 phenylpyruvic acid. The explanation for this is probably that both branched chain transaminase and aromatic amino acid transminase are feedback inhibited by L-phenylalanine, whereas aspartate phenylalanine transaminase is not inhibited by L-phenylalanine. In addition, since oxaloacetate, which is produced when aspartic add is used as the amino donor, is readily converted to pyruvic add, no feedback inhibition involving oxaloacetate occurs. The reason for low conversion yield of some E. coli strains might be that these E. cdi strains are defident in the aspartate phenylalanine transaminase. 

The best results were obtained with L-aspartic add as the amino donor for P. denitrificam and phenylpyruvic add as the amino acceptor. With L-aspartic add, conversion of phenylpyruvic add exceeded 90%. This may be attributed to absence of feedback inhibition of the reaction due to metabolism of file reaction product, oxaloacetic add. When using glutamic acid the conversion of phenylpyruvic add did not exceed 60%. 

Enzymes in the pathway to L-phenylalanine are subject to feedback inhibition by products (amino adds) arising from pathway intermediates. 

Auxotrophic mutant lack one or more enzymes involved in the synthesis of amino acids (such as tyrosine). This prevents accumulation of the amino acid and thus avoids feedback inhibition of enzymatic steps in the L-phenylalanine pathway. 

Regulatory mutants are not subject to feedback inhibition, even by L-phenylalanine itself. 

A Try mutant would not be subject to feedback inhibition by overproduction of tryptophan. Also, the mutation may allow more chorismate to proceed to prephenate via E3 (see Figure 8.4) and thus through to L-phenylalanine. 

At low doses, both psychostimulants could theoretically stimulate tonic, extracellular levels of monoamines, and the small increase in steady state levels would produce feedback inhibition of further release by stimulating presynaptic autoreceptors. While this mechanism is clearly an important one for the normal regulation of monoamine neurotransmission, there is no direct evidence to support the notion that the doses used clinically to treat ADHD are low enough to have primarily presynaptic effects. However, alterations in phasic dopamine release could produce net reductions in dopamine release under putatively altered tonic dopaminergic conditions that might occur in ADHD and that might explain the beneficial effects of methylphenidate in ADHD. 

Feedback inhibition refers to inhibition of an enzyme in a biosynthetic pathway by an end product of that pathway. For example, for the biosynthesis of D from A catalyzed by enzymes EnZj through Enz3,... 

In a branched biosynthetic pathway, the initial reactions participate in the synthesis of sevetal products. Figure 9—4 shows a hypothetical btanched biosynthetic pathway in which cutved attows lead from feedback inhibitors to the enTymes whose activity they inhibit. The sequences S3 —> A, S4 —> B, S4 —> C, and S3 — > D each represent hneat teaction sequences that are feedback-inhibited by theit end products. The pathways of nucleotide biosynthesis (Chaptet 34) provide specific examples. 

The kinetics of feedback inhibition may be competitive, noncompetitive, pattially competitive, ot mixed. Feedback inhibitots, which frequently ate the small molecule building blocks of mactomolecules (eg, amino acids for proteins, nucleotides fot nucleic acids), typically inhibit the fitst committed step in a particulat biosynthetic sequence. A much-studied example is inhibition of bacterial aspattate ttanscatbamoylase by CTP (see below and Chaptet 34). 

Multiple feedback loops can provide additional fine control. For example, as shown in Figure 9—5, the presence of excess product B decteases the tequitement for substrate 3. Howevet, Sj is also tequited fot synthesis of A, C, and D. Excess B should thetefote also curtail synthesis of all font end products. To circumvent this potential difficulty, each end product typically only partially inhibits catalytic activity. The effect of an excess of two or more end products may be strictly additive or, alternatively, may be greater than their individual effect (cooperative feedback inhibition). 

Aspartate transcarbamoylase (ATCase), the catalyst for the first reaction unique to pyrimidine biosynthesis (Figure 34-7), is feedback-inhibited by cytidine tri-... 

Figure 9-4. Sites of feedback inhibition in a branched biosynthetic pathway. Si-Sj are intermediates in the biosynthesis of end products A-D. Straight arrows represent enzymes catalyzing the indicated conversions. Curved arrows represent feedback loops and indicate sites of feedback inhibition by specific end products.

Figure 9-5. Multiple feedback inhibition in a branched biosynthetic pathway. Superimposed on simple feedback loops (dashed, curved arrows) are multiple feedback loops (solid, curved arrows) that regulate enzymes common to biosynthesis of several end products.

FEEDBACK REGULATION IS NOT SYNONYMOUS WITH FEEDBACK INHIBITION... 

Acetyl-CoA carboxylase is an allosteric enzyme and is activated by citrate, which increases in concentration in the well-fed state and is an indicator of a plentiful supply of acetyl-CoA. Citrate converts the enzyme from an inactive dimer to an active polymeric form, having a molecular mass of several milhon. Inactivation is promoted by phosphorylation of the enzyme and by long-chain acyl-CoA molecules, an example of negative feedback inhibition by a product of a reaction. Thus, if acyl-CoA accumulates because it is not esterified quickly enough or because of increased lipolysis or an influx of free fatty acids into the tissue, it will automatically reduce the synthesis of new fatty acid. Acyl-CoA may also inhibit the mitochondrial tricarboxylate transporter, thus preventing activation of the enzyme by egress of citrate from the mitochondria into the cytosol. 

Since biosynthesis of IMP consumes glycine, glutamine, tetrahydrofolate derivatives, aspartate, and ATP, it is advantageous to regulate purine biosynthesis. The major determinant of the rate of de novo purine nucleotide biosynthesis is the concentration of PRPP, whose pool size depends on its rates of synthesis, utilization, and degradation. The rate of PRPP synthesis depends on the availabihty of ribose 5-phosphate and on the activity of PRPP synthase, an enzyme sensitive to feedback inhibition by AMP, ADP, GMP, and GDP. 

Purine and pyrimidine biosynthesis parallel one another mole for mole, suggesting coordinated control of their biosynthesis. Several sites of cross-regulation characterize purine and pyrimidine nucleotide biosynthesis. The PRPP synthase reaction (reaction 1, Figure 34-2), which forms a precursor essential for both processes, is feedback-inhibited by both purine and pyrimidine nucleotides. 

Hepatic purine nucleotide biosynthesis is stringently regulated by the pool size of PRPP and by feedback inhibition of PRPP-glutamyl amidotransferase by AMP and GMP. 

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