Organophosphorus compounds enzyme inhibitors

Another different class of inhibitors binds covalently to specific amino acids in the enzyme and these are referred to as irreversible inhibitors. The organophosphorus compounds, of which nerve gases are examples, inactivate enzymes which rely on the hydroxyl group of serine residues for their activity, e.g. cholinesterase (EC 3.1.1.8). 

Irreversible inhibition in an organism usually results in a toxic effect. Examples of this type of inhibitor are the organophosphorus compounds that interfere with acetylcholinesterase (see Box 7.26). The organophosphorus derivative reacts with the enzyme in the normal way, but the phosphory-lated intermediate produced is resistant to normal hydrolysis and is not released from the enzyme. 

Selected entries from Methods in Enzymology [vol, page(s)] Types of organophosphorus inhibitors, 11,686-688 toxicity hazards, 11,688 purity and analysis, 11,688 solutions of organophosphorus compounds, 11,689 estimation of specific radioactivity of organophosphorus compounds, 11,689-690 method for estimating phosphorus content, 11,691 reactions with enzymes, 11,691 -701 [rate constants, 11,692 phosphorylation of chymotrypsin, 11, 694-696 identification of phosphoryl and phosphonyl peptides,... 

Other important enzyme inhibitors of this type are the organophosphorus compounds. Thus, after metabolism to the oxygen analogues, the insecticides parathion and malathion (chap. 4, Fig. 25) (Fig. 5.12) form complexes with the enzyme acetylcholinesterase as described in more detail in chapter 7. 

Inhibition of the cholinesterase enzymes depends on blockade of the active site of the enzyme, specifically the site that binds the ester portion of acetylcholine (Fig. 7.48). The organophosphorus compound is thus a pseudosubstrate. However, in the case of some compounds such as the phosphorothionates (parathion and malathion, for example), metabolism is necessary to produce the inhibitor. 

Among the irreversible inhibitors are organophosphorus compounds, which inhibit the enzyme acetylcholinesterase and similar enzymes. Organophosphorous compounds include nerve gases (such as sarin), that work on the human nervous system, and insecticides like malathion. 

Inhibitors are classified into reversible and irreversible types. Reversible inhibition implies that the activity of the enzyme is fiiUy restored when the inhibitor is removed from the system in which the enzyme acts by some physical separative process, such as dialysis, gel filtration, or chromatography. An irreversible inhibitor combines covalently with the enzyme so that physical methods are ineffective in separating the two. For example, organophosphorus compounds are... 

The commercial use of some organophosphates and carbamates as systemic insecticides has resulted in the synthesis of a large number of these anticholinesterases. Structural formulae and some data for these inhibitors are given in the reviews of Usdin (Ul), Aldridge and Reiner (A12), and Main (M4). Compounds which structurally resemble substrates are usually very good inhibitors of plasma cholinesterase. Thus, organophosphorus compounds modeled on acetylcholine are powerful inhibitors of the enzyme (Table 20). Quaternary aminophenylphosphates... 

Organophosphorus compounds are irreversible inhibitors of acetylcholine esterase and butyrylcholine esterase (BuChe, EC 3.1.1.8) because the phosphate group is irreversibly bound by the enzyme. Therefore, organophosphorus pesticides can be detected by using the free enzyme. Since the activity of cholinesterases (ChE) in normal serum is rather large (800 UA), untreated serum pools may be employed for inhibitor determination. Gruss and Scheller (1987) have shown that the hydrolysis of butyrylthiocholine iodide can be directly indicated at a membrane-covered platinum electrode polarized to +470 mV. Twenty seconds after sample addition a steady value proportional to the enzyme activity was obtained in the differentiated current-time curve. Injection of an inhibitor decreased the rate of thiocholine formation, so that the residual activity could be evaluated after 30 s (Fig. 115). 

An example of absolutely cumulative toxicity is afforded by tri-o-cresyl phosphate or TOCP (figure 2,9). This compound is a cholinesterase inhibitor and neurotoxin. In chickens, an acute dose of 30 mg/kg has a severe toxic effect, which is produced to the same extent by a dose of 1 (mg/kg)/day given for 30 days. This effect may of course be produced by accumulation of the compound in vivo to a threshold toxic level, or it may result from the accumulation of the effect, as it probably does in the case of TOCP. Thus, the inhibition of cholinesterase enzymes by organophosphorus compounds may... 

Other types of inhibitors may not be so tolerable. Organophosphorus compounds, used in nerve gases and weed killers (e.g., parathion), form a covalent irreversible bond with the active serine and permanently inactivate acetylcholinesterase. This is a type of suicide inhibition because the inhibitor reacts with the enzyme much like a substrate, but becomes blocked in the intermediate state where the enzyme-phosphoryl bond is stable, in contrast to the hydrolyzable enzyme-acetyl bond. These compounds are life-threatening. 

The nerve gases are powerful inhibitors of the enzyme acetylcholinesterase (AChE), which causes rapid hydrolysis of acetylcholine. They are also inhibitors of butyryl-choUnesterase and carboxylesterase. Acetylcholine is the chemical that is released to transmit nerve impulses in the central nervous system and also at several peripheral locations. Once the impulse is transmitted, acetylcholine must be removed instantaneously for proper functioning of the nervous system. Such removal is done by the enzyme AChE, which is found extensively in nervous system and many non-nervous tissues. Nerve gases and other organophosphorus compounds bind... 

A number of important compounds have the ability to combine with certain enzymes and prevent their subsequent reaction with substrates. Such compounds are called enzyme inhibitors. Many drugs and poisons act by enzyme inhibition. Organophosphorus insecticides and nerve gases fall into this category. Immobilised enzymes, which have applications in industrial technology, are enzymes which have been processed to increase their stability so that they can be repeatedly used (Figure 11.14). 

Organophosphorus compounds are widely used in agriculture as insecticides, in industry and in military technology as chemical warfare agents. They are extremely potent inhibitors of the enzyme AChE, which is responsible for the termination of the action of acetylcholine (ACh) at cholinergic synapses. 

The presence of low concentrations of inhibitors strongly and specifically affects enzyme activity. Therefore, by measuring the enzyme activity the concentration of the organophosphorus compounds can be assayed. 

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