Diisopropyl fluorophosphate toxicity

The organophosphorus cholinesterase inhibitors like diisopropyl fluorophosphate, phospholine, parathion, malathion etc. are highly toxic compound and cause irreversible inhibition of both true and pseudocholinesterases. They are highly lipid soluble compound and can easily cross the blood-brain barrier. 

The serine proteases act by forming and hydrolyzing an ester on a serine residue. This was initially established using the nerve gas diisopropyl fluorophosphate, which inactivates serine proteases as well as acetylcholinesterase. It is a very potent inhibitor (it essentially binds in a 1 1 stoichiometry and thus can be used to titrate the active sites) and is extremely toxic in even low amounts. Careful acid or enzymatic hydrolysis (see Section 9.3.6.) of the inactivated enzyme yielded O-phosphoserine, and the serine was identified as residue 195 in the sequence. Chy-motrypsin acts on the compound cinnamoylimidazole, producing an acyl intermediate called cinnamoyl-enzyme which hydrolyzes slowly. This fact was exploited in an active-site titration (see Section 9.2.5.). Cinnamoyl-CT features a spectrum similar to that of the model compound O-cinnamoylserine, on denaturation of the enzyme in urea the spectrum was identical to that of O-acetylserine. Serine proteases act on both esters and amides. 

Grigoryan, H., Schopfer, L.M., Thompson, C.M., Terry, A.V., Masson, P., Lockridge, O. (2008). Mass spectrometry identifies covalent binding of soman, sarin, chlorpyrifos oxon, diisopropyl fluorophosphate, and FP-biotin to tyrosines on tubulin a potential mechanism of long term toxicity by organophos-phoms agents. Chem. Biol. Interact. April 22. (Epub ahead of print)... 

Stone, J.D., Terry, A.V., Pauly, J.R., Trendergast, M.A., Buccafusco, J.J. (2000). Protractive effects of chronic treatment with an acutely suh-toxic regimen of diisopropyl-fluorophosphate on the expression of cholinergic receptor densities in rats. Brain Res. 882 9-18. 

Most irreversible enzyme inhibitors combine covalently with functional groups at the active sites of enzymes. These inhibitors are usually chemically reactive, and many of them show some specificity in terms of the amino acid groups which they react with. Diisopropyl fluorophosphate (DFP), for example, forms a covalent adduct with active site serine residues, such as in the serine proteases, and in acetylcholinesterase, which explains its toxic effect on animals. Irreversible enzyme inhibition can be used to identify important active site residues. A special case of irreversible enzyme inhibition is the effect of suicide inhibitors, which are generally chemically unreactive compounds that resemble the substrate of the target enzyme and bind at the active site. The process of enzyme turnover begins, but the inhibitor is so... 

Fluorophosphates are also highly toxic and relatively volatile. Sarin and soman are chemical warfare agents. Diisopropyl fluorophosphate (DFP) is often used by biochemists to study serine-active enzymes. Mipafox and DFP cause OPIDN in humans and experimental animals. 

The effect in vivo of diisopropyl fluorophosphate upon the plasma of several adults has been measured (M22). A distinct fall in enzymic activity was observed, but no clinical signs of intoxication appeared in any individual. The half-life of the cholinesterase was found to be 16 days, thus confirming that alterations in the plasma enzyme activity are of limited value in assessing the toxic effects of diisopropyl fluorophosphate. 

Acetylcholinesterase, the enzyme that hydrolyzes acetylcholine to choline in the postsynaptic membrane, is a serine esterase inhibited by diisopropyl fluorophosphate, sarin, physostigmine, and parathion (Table 11.4). These substances are extremely toxic and cause paralysis. Other toxins block the acetylcholine receptor (antagonists) or lock it open (agonists). Nicotine is an agonist. 

Such inhibition involves covalent bonding at the active site and cannot be reversed by excess substrate or by dialysis. The site is therefore blocked and made catalytically inactive. Most inhibitors in this group are highly toxic, e.g. the organophosphorus nerve poisons. Thus, diisopropyl fluorophosphate (DFP) reacts irreversibly with the hydroxyl group of serine ... 

Diisopropyl fluorophosphate (DFP), a military nerve gas used during World War II, inactivates acetylcholinesterase by reacting with its CH2OH group. When the enzyme is inactivated, nerve impulses cannot be transmitted properly and paralysis occurs. DFP is extremely toxic. Its LD50 (the lethal dose for 50% of the test animals) is only 0.5 mg/kg of body weight (page 922). 

In the balance of this paper, we describe the application of our method of chemical modification to the generation of fluorohydrolases The most common substrate to measure fluorohydrolase activity is diiso-propylphosphorofluoridate (DFP). Although DFP is not found in nature, Mazur reported the enzymatic hydrolysis of this highly toxic organo-fluorophosphate by an enzyme isolated from hog kidney.Since then, diisopropyl phosphorofluoridate fluorohydrolase (DFPase, E.C. 3.8.2.1) has been extensively studied and well characterized. In addition, enzymes with organofluorophosphate hydrolyzing activity have been shown to be widely distributed phylogenetically and sources are known from bacteria, protozoa, invertebrates, and vertebrates. 

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