Malonate succinate dehydrogenase inhibition

The enzyme succinate dehydrogenase (SDH) is competitively inhibited by malo-nate. Figure 14.14 shows the structures of succinate and malonate. The structural similarity between them is obvious and is the basis of malonate s ability to mimic succinate and bind at the active site of SDH. However, unlike succinate, which is oxidized by SDH to form fumarate, malonate cannot lose two hydrogens consequently, it is unreactive. 

Barbiturates such as amobarbital inhibit NAD-hnked dehydrogenases by blocking the transfer from FeS to Q. At sufficient dosage, they are fatal in vivo. Antin cin A and dimercaprol inhibit the respiratory chain between cytochrome b and cytochrome c. The classic poisons H2S, carbon monoxide, and cyanide inhibit cytochrome oxidase and can therefore totally arrest respiration. Malonate is a competitive inhibitor of succinate dehydrogenase. 

The activity of complex II (succinate dehydrogenase) is measured as the succinate-dependent reduction of decylubiquinone, which is in turn recorded spectro-photometrically through the reduction of dichlorophenol indophenol at 600 nm (e 19,100-M -cm Fig. 3.8.5). In order to ensure a linear rate for the activity, the medium is added with rotenone, ATP, and a high concentration of succinate. As noticed previously for complex I, decylubiquinone is not a perfect acceptor for electrons from the membrane-inserted complex II [70]. Malonate, a competitive inhibitor of the enzyme, is used to inhibit it. Rather than decylubiquinone, phenazine methosulfate can be utilized, which diverts the electrons from the complex before they are conveyed through subunits C and D, therefore allowing measurement of the activity of subunits A and B. 

Competitive inhibitors often closely resemble in some respect the substrate whose reactions they inhibit and, because of this structural similarity, compete for the same binding site on the enzyme. The enzyme-inhibitor complex either lacks the appropriate reactive groups or is held in an unsuitable position with respect to the catalytic site of the enzyme which results in a complex which does not react (i.e. gives a dead-end complex). The inhibitor must first dissociate before the true substrate may enter the enzyme and the reaction can take place. An example is malonate, which is a competitive inhibitor of the reaction catalysed by succinate dehydrogenase. Malonate has two carboxyl groups, like the substrate, and can fill the substrate binding site on the enzyme. The subsequent reaction, however, requires that the molecule be reduced with the formation of a double bond. If malonate is the substrate, this cannot be achieved without the loss of one of the carboxy-groups and therefore no reaction occurs. 

Figure 7.3 The similarity of the structures of malonate and succinate explains why malonate inhibits succinate dehydrogenase...

The classic example of competitive inhibition is inhibition of succinate dehydrogenase, an enzyme, by the compound malonate. Hans Krebs first elucidated the details of the citric acid cycle by adding malonate to minced pigeon muscle tissue and observing which intermediates accumulated after incubation of the mixture with various substrates. The structure of malonate is very similar to that of succinate (see Figure 1). The enzyme will bind malonate but cannot act further on it. That is, the enzyme and inhibitor form a nonproductive complex. We call this competitive inhibition, as succinate and malonate appear to compete for the same site on the enzyme. With competitive inhibition, the percent of inhibition is a function of the ratio between inhibitor and substrate, not the absolute concentration of inhibitor. 

Other researchers had found that arsenate (AsOf ) inhibits a-ketoglutarate dehydrogenase and that malonate inhibits succinate dehydrogenase. 

A classic example of competitive inhibition is the inhibition of succinate dehydrogenase by malonate, a structural analogue of succinate. Competitive inhibitors are usually structural analogues of the substrate, the molecule with which they are competing. They bind to the active site but either do not have a structure that is conducive to enzymatic modification or do not induce the proper orientation of catalytic amino acyl residues required to affect catalysis. Consequently, they displace the substrate from the active site and thereby depress the velocity of the reaction. Increasing [S] will displace the inhibitor. 

There are two Krebs cycle inhibitors that are worth mentioning. Malonate inhibits succinate dehydrogenase because of its very similar structure. Fluoro-acetate inhibits cis-aconitase, which is an Fe-S enzyme. The fluoroacetate replaces acetate as a substrate in the citrate synthase reaction when this combines with cis-aconitase, however, no further reaction becomes possible. 

The addition of malonate, a specific inhibitor of the succinic dehydrogenase system, inhibits the regeneration of oxalacetic acid and results in the accumulation of citric, a-ketoglutaric and succinic acids. Upon exhaustion of the remaining stores of oxalacetic acid, pyruvate oxidation in muscle ceases and can only be resumed after further addition of oxalacetic acid or addition of any dicarboxylic acid capable of conversion to oxalacetic acid. Under these conditions, the amount of pyruvate oxidized is equivalent to the amount of oxalacetate supplied. 

Malonic acid is a classical example of a true competitive inhibitor. Malonic acid inhibits succinic dehydrogenase, which catalyzes the oxidation of succinic acid to fumaric acid, as shown below. 

Succinate dehydrogenase is competitively inhibited by malonate, the next lower homologue of succinate (Chapter 6). 

During the terminal stages of electron transfer in complex II, cytochrome bysg is involved however, its specific function is not understood. Oxaloacetate and malonate are competitive inhibitors of succinate dehydrogenase and compete with the substrate for binding at the active site (Chapters 6 and 13). Carboxin and thenoyltrifluoroacetone (Figure 14-7) inhibit electron transfer from FADH2 to CoQ. 

Having a lifetime of two weeks, the sensor was capable of determining NADH and succinate with a linear range up to 0.13 and 0.15 mmolA, respectively. Competitive inhibition of the mitochondrial succinate dehydrogenase activity by malonate resulted in a totally selective sensor for NADH.In this way the selectivity enhancement of HIS sensors by inhibition of interfering metabolic routes was demonstrated. 

In addition, feedback inhibition has two other characteristics that would be unlikely to arise haphazardly in longer pathways there is often very little structural resemblance between the end product that inhibits and the substrate of the inhibited enzyme and the inhibition is usually cooperative. Let us consider what these two properties mean. When inhibition arises for no obvious biological reason, it is usually for an obvious enough chemical reason the substrate and inhibitor are similar enough in terms of chemical structure that the inhibitor can bind to the same site on the enzyme as the one where the substrate binds however, as it lacks some feature necessary for the chemical reaction it does not react but does nothing. A classical example is provided by the enzyme succinate dehydrogenase, which uses succinate as its substrate but is inhibited by malonate (Figure 10.3). Succinate and malonate have almost the same chemical structures, so either is likely to bind to a site intended for... 

Although enzymes catalyze only certain reactions or certain types of reaction, they are still subject to interference. When the activated complex is formed, the substrate is adsorbed at an active site on the enzyme. Other substances of similar size and shape may be adsorbed at the active site. Although adsorbed, they will not undergo any transformation. However, they do compete with the substrate for the active sites and slow down the rate of the catalyzed reaction. This is called competitive inhibition. For example, the enzyme succinic dehydrogenase will specifically catalyze the dehydrogenation of succinic acid to form fumaric acid. But other compounds similar to succinic acid will competitively inhibit the reaction. Examples are other diprotic acids such as malonic and oxalic acids. Competitive inhibition can be reduced by increasing the concentration of the substrate relative to that of the interferent so that the majority of enzyme molecules combine with the substrate. 

The enzyme is unable to bind both S and I at the same time and in competitive inhibition, the enzyme-inhibitor complex El does not react with substrate S. Competitive inhibitors often resemble substrate structurally. As an example we can mention malonate, which is an inhibitor for dehydrogenation of succinate of a enzyme succinic dehydrogenase and resembles the structure of succinate (Figure 6.35)... 

Addition of malonate, a specific inhibitor for succinate dehydrogenase, or antlmycln-A, which blocks the respiratory electron transfer chain between cyt-b and cyt-cj, resulted In severe Inhibition of metal complexatlon (Figure 11), This Implies that the actual m.etal reductant occurs subsequent to cyt-b in the electron-transfer sequence and either cyt-c or cyt-c are likely candidates. Assuming that either cytochrome serves to reduce Ru(iri) at least partially explains the low yield of the reaction, since the reduction potentials of these proteins CO.225 and 0.254 V, respectively) are rather high relative to that of the metal complex 0-0.042 V) and sterlc interactions would probably prevent close contact with the reducing heme moiety C40-41). 

The next relevant discovery was made in 1910, when it was noted that some enzymes are blocked by substances whose molecular structure resembles that of the normal substrates. Thus amylase, which normally hydrolyses starch, is inhibited by glucose (see Section 9.1). Again, malonic acid 9.3) competitively inactivates the enzyme succinic dehydrogenase by displacing the normal substrate, succinic acid 9.4), from the enzyme (Quastel and Wooldridge, 1927). A similar phenomenon in physiology is the toxic action of carbon monoxide... 

A competitive inhibitor binds to the active site of an enzyme and thus competes with snbstrate molecnles for the active site. Competitive inhibitors often have molecular structures that are similar to the normal substrate of the enzyme. The competitive inhibition of snccinate dehydrogenase by malonate is a classic example. Succinate dehydrogenase catalyzes the oxidation of the substrate succinate to form fumarate by transferring two hydrogens to the coenzyme FAD ... 

Malonate, having a structure similar to succinate, competes for the active site of succinate dehydrogenase and thus inhibits the enzyme ... 

The best-known example for the competitive inhibition shown in reaction (5.1) is the inhibition of succinate dehydrogenase by malonate, a compound stmcturaUy closely related to succinate (Price Stewens, 1999). 

An example of competitive inhibition is the inhibition of succinate dehydrogenase by malonate ion. The catalyzed reaction is the oxidation of succinate ion to fumarate ion ... 

Allen et al., 1964), where they are known to catalyze the conversion of oxaloacetate to succinate at a high rate (Krebs and Egglestone, 1941). In propionibacteria, the succinate dehydrogenase is not inhibited by malonate, in contrast with succinate dehydrogenases of the Krebs cycle (Ichikawa, 1955). 

The presence of I reduces the amount of E-S formed but, if more substrate is added, I eompetes less effectively and the inhibition may be overcome. A notable example of this tjrpe of inhibition is the effect of malonate on succinate dehydrogenase which catalyses the reaction... 

Hydrogen atoms are transferred to enzyme-bound FAD, and hence to ubiquinone and the respiratory chain. Succinate dehydrogenase is competitively inhibited by the homologue of succinate, malonate. Only the /rans-isomer fumarate is generated. 

The Oxaloacetic System (Szent-Gyoi i, 1937).— The wide distribution of the three enzymes, succinic and malic dehydrogenase, and fumarase, si ests that they and their respective substrates participate in many tissue respirations. Addition of malonic acid, which inhibits succinic acid oxidation, leads to an almost complete... 

The efficiency of an enzyme can be reduced or can even become negligible in the presence of certain substances, known as inhibitors. Many inhibitors have structural resemblances with the substrates and compete with them for the formation of complexes with the enzyme. This is the case of the inactivation of cytochrome c oxidase by the cyanide ion, which blocks the mitochondrial electron-transport chain to oxygen. Similarly, the inactivation of the succinate dehydrogenase by malonate involves its inhibition of the conversion of succinate to fumarate in the citric acid cycle. In the latter case, the mechanism for competitive inhibition is... 

Competitive inhibition occurs whenever the place on the enzyme which should be occupied by the substrate molecule is taken by another molecule which cannot itself react. The inhibiting molecule blocks the enzyme. One clasacal example is the pair succinate (substrate) and malonate (inhibitor) with succinate dehydrogenase. Raising the substrate concentration replaces the inhibitor (malonate) on the surface of the enzyme according to the law of mass action. Mathematical analysis of the extent of inhibition as a fimction of inhibitor and substrate concentrations can easily distinguish between competitive and noncompetitive inhibition. 

Dehydrogenation of succinate introduces a C—C double bond to give rise to the frans-compound fumarate (step 7). The enzyme succinate dehydrogenase is a flavoprotein and is a member of the respiratory chain (cf. Chapt. X-4). This reaction is inhibited by malonate—the classical example of competitive inhibition The wrong substrate is attached to the enzyme, but cannot undergo the reaction for simple chemical reasons. 

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