Enzyme reversible inhibition

The three reversible mechanisms for enzyme inhibition are distinguished by observing how changing the inhibitor s concentration affects the relationship between the rate of reaction and the concentration of substrate. As shown in figure 13.13, when kinetic data are displayed as a Lineweaver-Burk plot, it is possible to determine which mechanism is in effect. 

Following concurrent administration of two drugs, especially when they are metabolized by the same enzyme in the liver or small intestine, the metabolism of one or both drugs can be inhibited, which may lead to elevated plasma concentrations of the dtug(s), and increased pharmacological effects. The types of enzyme inhibition include reversible inhibition, such as competitive or non-competitive inhibition, and irreversible inhibition, such as mechanism-based inhibition. The clinically important examples of drug interactions involving the inhibition of metabolic enzymes are listed in Table 1 [1,4]. 

Figure 5.8 Dilution scheme for testing the reversibility of an enzyme inhibition The enzyme and inhibitor are pre-incubated at a concentration of enzyme equal to 100-fold that needed for activity assay, and at a concentration of inhibitor equal to 10-fold the IC50 value. The sample is then rapidly diluted 100-fold into an assay solution. The inhibitor concentration thus goes from 10-fold above die IC50 (corresponding to 91% inhibition) to 10-fold below the IC50 (corresponding to 9% inhibition).

Until now our discussions of enzyme inhibition have dealt with compounds that interact with binding pockets on the enzyme molecule through reversible forces. Hence inhibition by these compounds is always reversed by dissociation of the inhibitor from the binary enzyme-inhibitor complex. Even for very tight binding inhibitors, the interactions that stabilize the enzyme-inhibitor complex are mediated by reversible forces, and therefore the El complex has some, nonzero rate of dissociation—even if this rate is too slow to be experimentally measured. In this chapter we turn our attention to compounds that interact with an enzyme molecule in such a way as to permanendy ablate enzyme function. We refer to such compounds as enzyme inactivators to stress the mechanistic distinctions between these molecules and reversible enzyme inhibitors. 

Zinc is in the active site of ALAD and can play a protective role in lead intoxication by reversing the enzyme-inhibiting effects of lead. Children with high PbB levels (50-67 pg/dL) were reported to consume less zinc than children with lower PbB (12-29 pg/dL) (Johnson and Tenuta 1979). In a group of 13 children, Markowitz and Rosen (1981) reported that the mean serum zinc levels in children with... 

Because of the complexity of biological systems, Eq. (1) as the differential form of Michaelis-Menten kinetics is often analyzed using the initial rate method. Due to the restriction of the initial range of conversion, unwanted influences such as reversible product formation, effects due to enzyme inhibition, or side reactions are reduced to a minimum. The major disadvantage of this procedure is that a relatively large number of experiments must be conducted in order to determine the desired rate constants. 

Reversible enzyme inhibition, 10 256, 318 Reversible heterogeneous catalytic reactions, 25 312 Reversible hydrogenase-based biophotolysis, 13 849 Reversible perturbation reactions, 14 617 Reversible process, 24 650 Reversible superconducting region, 23 818 Reversible toxicity, 25 203 Revert titanium, recycling of, 24 848, 857 Review activities, EIA, 10 237 Rexfoam, commercial defoamer, 8 241t Rexillium III... 

It will be shown below that D.F.P. is rapidly destroyed in vitro and in vivo.2 Therefore, the recovery of serum cholinesterase activity is not representative of a reversal of enzyme inhibition, but is indicative of synthesis of new enzyme proteins. Since the regeneration rate of serum cholinesterase in patients with liver damage is significantly depressed as contrasted with that in the normal patient, it is concluded that the ability of such patients to synthesize this particular enzyme protein is decreased. This constitutes evidence for the view that the fiver is a primary locus for the formation of serum cholinesterase. 

The mechanisms which underlie enzyme inhibition are described more fully in Chapter 3. Suffice to say here that reversible inhibitors which block the active site are called competitive whilst those which prevent release of the product of the reaction are non-competitive. By preventing the true substrate accessing the active site, competitive inhibitors increase Km (designated by or K PParent). A non-competitive inhibitor decreases V mprime symbol ( ) here to imply physiological as it does for energy change. 

The problem with enzyme catalyzed reversible transesterifications as an approach to biochemical resolution is that due to the reversible nature of the reaction, the enantiomeric excess of the desired product in the forward reaction decreases as the reverse reaction proceeds. As in hydrolytic reactions, the irreversible transesterification offers a better process for optimization of the transformation and for recovery of the product. Furthermore, there is no product inhibition observed in the irreversible process. Both enantiomers can be converted to glycerol acetonide with known procedures. 

Qiolinesterases from various sources Quaternary heterocyclic oximes inhibit enzyme activity reversibly 127-131... 

Antibodies against the virus but also amantadine and derivatives, interfere with host cell penetration. There are nucleoside analogues such as aciclovir and ganciclovir, which interfere with DNA synthesis, especially of herpes viruses. Others like zidovudine and didanosine, inhibit reverse transcriptase of retroviruses. Recently a number of non-nucleoside reverse transcriptase inhibitors was developed for the treatment of HIV infections. Foscarnet, a pyrophosphate analogue, inhibits both reverse transcriptase and DNA synthesis. Protease inhibitors, also developed for the treatment of HIV infections, are active during the fifth step of virus replication. They prevent viral replication by inhibiting the activity of HIV-1 protease, an enzyme used by the viruses to cleave nascent proteins for final assembly of new vi-rons. 

All NSAIDs except aspirin inhibit cyclooxygenase reversibly. Inhibition by aspirin, caused by the covalent acetylation of the enzyme, is irreversible. In platelets most NSAIDs block thromboxane synthesis more than that of prostacyclin and the overall effect is therefore inhibition of platelet aggregation. This effect is already noticeable at low doses. Because of the irreversible nature of the enzyme inhibition by aspirin and the fact that in platelets the novo enzyme synthesis is not possible the aggregation inhibitory effects of aspirin last several days. 

Another important characteristic of M AOIs is the production of reversible versus irreversible enzyme inhibition. An irreversible inhibitor permanently disables the enzyme. This means that MAO must be resynthesized, in the absence of the drug, before the activity of the enzyme can be reestablished. Resynthesis of the enzyme may take up to 2 weeks. For this reason, an interval of 10-14 days is required after discontinuing irreversible inhibitors and before instituting treatment with other antidepressants or permitting the use of contraindicated drugs or the consumption of contraindicated foods. On the other hand, a reversible inhibitor can move away from the active site of the enzyme, making the enzyme available to metabo-hze other substances. The reversibility and selectivity of the currently available MAOIs are summarized in Table 2-4. 

Enzyme inhibitors inhibit the action of enzymes either reversibly or irreversibly. Since enzymes are such pervasive, powerful biological catalysts, inhibitors can act as potent drugs. Broadly categorized, enzyme inhibitors may be either irreversible or reversible. 

The hormone-hke peptide incretin stimulates the release of insuhn by a feedback process that involves cleaving the molecule to an inactive form. The protease enzyme dipeptidal peptidase (DPP) in turn cleaves incretin, in effect inactivating this enzyme. Inhibition of DPP consequently extends the action of incretin. This inhibition thus prevents the increased levels of blood glucose that mark diabetes. The protease inhibitor vidagliptin, which is modeled in part on the terminal sequence in DPP, has been found to sustain levels of insulin in Type II diabetics. The inhibition is apparently reversible in spite of the presence in the structure of the relatively reactive a-aminonitrile function. Construction of one intermediate in the convergent synthesis comprises the reaction of amino adamantamine (21-1) with a mixture of nitric and... 

We have studied the molecular mechanism of the enzyme inhibition using bovine erythrocyte AChE 14). Depending upon the position and nature of substituents, the value of kt/k i (= 1/Kd) showed significant variations whereas that of k2 did not. Thus, formation of the reversible complex was considered to be the step which governs the variation in overall inhibitory activity. 

The process of reversible inhibition is described by an equilibrium interaction between enzyme and inhibitor. Most inhibition processes can be classified as competitive or noncompetitive, depending on how the inhibitor impairs enzyme action. A competitive inhibitor is usually similar in structure to the substrate and is capable of reversible binding to the enzyme active site. In contrast to the substrate molecule, the inhibitor molecule cannot undergo chemical transformation to a product however, it does interfere with substrate binding. A noncompetitive inhibitor does not bind in the active site of an enzyme but binds at some other region of the enzyme molecule. Upon binding of the noncompetitive inhibitor, the enzyme is reversibly converted to a nonfunctional conformational state, and the substrate, which is fully capable of binding to the active site, is not converted to product. 

Many amino acids are weak inhibitors of the various tissue phosphatases (26), and where investigated in more detail the inhibition has been found to be noncompetitive or mixed (178). The effects appear to vary considerably with the nature of the particular enzyme. Cysteine and histidine probably inhibit by virtue of their Zn2+ chelating ability (107, 179). Other compounds in this category include iodosobenzoate, iodo-acetamide (107), and Zn2+ (174). One component of urea inactivation of human tissue phosphatases has been shown to be a noncompetitive inhibition, reversible on dilution (53). 

Enzymatic techniques have also been employed in the analysis of these compounds. The toxicity of carbamate insecticides is due to the inhibition of the enzyme acetylcholine esterase, so the determination of these compounds can be achieved by enzyme inhibition (2,83,119), bioassay (118,167), or enzyme-linked immunosorbent assay (ELISA) (168-171). In the detection of carbamates by fluorimetric enzyme inhibition, the effluent from a reversed-phase chromatographic column was incubated with cholinesterase, which was introduced via a postcolumn reagent delivery pump. Then, the resulting partially inhibited cholinesterase was reacted with N-methyl indoyl acetate to produce a fluorophore and a reduction in the baseline fluorescence (172). 

FIGURE 2-50. The consequence of a substrate competing successfully for reversal of enzyme inhibition is that the substrate essentially displaces the inhibitor and shoves it off. Because the substrate has this capability, the inhibition is said to be reversible. 

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