The butyrylcholinesterases

Humans have several variants of BuChE, and as many as 11 forms of serum BuChEs have been described. One of these forms is associated with unusual clinical responses to succinylcholin, which is used as a muscle relaxant during surgery. Low-rate exposure of humans or other organisms to organophosphates or carbamates may cause an inhibition of the BuChE, without any sign of poisoning. 

An example may illustrate how sensitive plasma BuChE is to inhibition. Rats dosed orally with bromophos needed only 10.1 mg/kg of body weight for the enzyme activity to be reduced to half, whereas the erythrocyte AChE I50 was 1938 mg/kg and the brain AChE I50 was 576 mg/kg. The plasma 150 dose did not give any symptoms of poisoning (oral LD50 for rats = 3750 to 7700 mg/kg). It is very clear that plasma enzymes may be inhibited at much lower concentrations than needed to give the more severe symptoms associated with AChE inhibition in the nervous system, and they can be used in early-warning monitoring (Shivanandappa et al., 1988). 

The insecticides leptophos, EPN, cyanofenphos, trichloronate, and diox-abenzophos (salithion) cause irreversible ataxia not only to chickens but also to mice and sheep. It is believed that AChE inhibition is the reason for their acute toxicity, while NTE inhibition is responsible for causing paralytic ataxia. 

The esterase got the names neurotoxic esterase or neuropathy target esterase, and the ester phenyl valerate (PV) was found to be a good substrate for this esterase. However, PV is also hydrolyzed by other esterases because paraoxon, which does not give symptoms of delayed neurotoxicity, inhibited the PV activity as much as 80%. The NTE is defined as the hydrolytic activity against PV that is not inhibited by paraoxon but by mipafox. About 3% of the activity is not inhibited by mipafox plus paraoxon. Thus, the part of PV activity due to neurotoxic esterase should be 17%. 

But this is not the whole story. It was soon established that many compounds could inhibit this particular activity without causing delayed neurotoxicity. In fact, many inhibitors protected the animals against established neurotoxic inhibitors if administered as pretreatment. 

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Stone et al. use this method to simultaneously synthesize the silica and entrap the butyrylcholinesterase which retains all its activity after the process of encapsulation, a high enzyme loading (90 %) is reached and the stability is increased [168]. The method has been further developed to simultaneously entrap catalase and horseradish peroxidase with inorganic magnetic nanopartides [169] which will fadlitate the separation [170,171]. 

In clinical diagnosis of OP exposure, the tissue most readily available for study is blood. OP adducts of butyrylcholinesterase are better candidates for study than OP adducts on acetylcholinesterase for the following reasons. Human blood contains 5 mg of butyrylcholinesterase and 0.5 mg of acetylcholinesterase per liter. The butyrylcholinesterase is in plasma, whereas the acetylcholinesterase is bound to the membranes of red and white cells. Most OPs, with the exception of chemical warfare nerve agents, react more rapidly with butyrylcholinesterase than with acetylcholinesterase. 

Yuan J, Yin J, Wang E (2007) Characterization of procaine metabolism as probe for the butyrylcholinesterase enzyme investigation by simultaneous determination of procaine and its metabolite using capillary electrophoresis with electrochemiluminescence detection. J Chromatogr A 1154(l-2) 368-372. doi 10.1016/j.chroma.2007.02.024... 

The ability of the electrode to detect choline over a wide concentration range allows its use for measurement of butyryl-choline concentration in assay systems employing coupled butyrylcholinesterase. The butyrylcholinesterase causes choline production due to enzymatic hydrolysis of butyryl-choline. 

Clayton and Purcell have reported excellent correlations using dipole moments, amide group moments, and Taft s polar substituent constants a ) along with hydrophobic parameters when applied to an homologous series of molecules.A regression of the butyrylcholinesterase inhibitory potencies of these mono(carbamoylpiperidino)decanes gave eq 5 in which... 

Podoly, E., Shalev, D.E., Shenhar-Tsarfaty, S., et al, 2009. The butyrylcholinesterase K variant confers structurally derived risks for Alzheimer pathology. J. Biol. Chem. 284 (25), 17170-17179. 

Galanthamine (23) is an alkaloid extracted from the common snowdrop Galanthus nivalis. This compound is a long-acting, competitive AChE inhibitor which appears to be somewhat more specific for acetylcholinesterase than plasma butyrylcholinesterase (132). It is well tolerated during long-term treatment (133) and is being evaluated clinically for AD (134). 

Metrifonate [52-68-6] (24) is itself not an AChE inhibitor, but is none2ymaticaIly converted into an active irreversible inhibitor of the en2yme. The compound is relatively specific for AChE over butyrylcholinesterase (135) and the irreversible nature of its inhibition gives rise to an extended duration of action. Some clinical experience has been gained through its use to treat schistosomiasis (136,137) and it is undergoing clinical evaluation for AD. 

There is a second type of cholinesterase called butyrylcholinesterase, pseudocholinesterase, or cholinesterase. This enzyme is present in some nonneural cells in the central and peripheral nervous systems as well as in plasma and serum, the liver, and other organs. Its physiologic function is not known, but is hypothesized to be the hydrolysis of esters ingested from plants (Lefkowitz et al. 1996). Plasma cholinesterases are also inhibited by organophosphate compounds through irreversible binding this binding can act as a detoxification mechanism as it affords some protection to acetylcholinesterase in the nervous system (Parkinson 1996 Taylor 1996). 

Acetylcholine, acetylcholinesterase, and butyrylcholinesterase are involved in the development of the nervous system (Brimijoin and Koeninsberger 1999 Layer 1990 Layer and Willbold 1994) some of this development is not complete until adulthood. Therefore, toxic chemicals acting on these substances could cause deleterious developmental effects in addition to the typical physiological effects already discussed. 

Hahn T, Ruhnke M, Luppa H. 1991. Inhibition of acetylcholinesterase and butyrylcholinesterase by the organophosphorus insecticide methyl parathion in the central nervous system of the golden hamster i Mesocricetus aumtus). Acta Histochem (Jena) 91 13-19. 

Cholinesterases are another group of B-esterases. The two main types are acetylcholinesterase (EC 3.1.1.7) and unspecific or butyrylcholinesterase (EC 3.1.1.8). Acetylcholinesterase (AChE) is found in the postsynaptic membrane of cholinergic... 

Kjellstrand P, Hohnquist B, Aim P, et al. 1983a. Trichloroethylene Further studies of the effects on body and organ weights and plasma butyrylcholinesterase activity in mice. Acta Pharmacol Toxicol 53 375-384. 

Released ACh is broken down by membrane-bound acetylcholinesterase, often called the true or specific cholinesterase to distinguish it from butyrylcholinesterase, a pseudo-or non-specific plasma cholinesterase. It is an extremely efficient enzyme with one molecule capable of dealing with something like 10000 molecules of ACh each second, which means a short life and rapid turnover (100 ps) for each molecule of ACh. It seems that about 50% of the choline freed by the hydrolysis of ACh is taken back into the nerve. There is a wide range of anticholinesterases which can be used to prolong and potentiate the action of ACh. Some of these, such as physostigmine, which can cross the blood-brain barrier to produce central effects and neostigmine, which does not readily... 

The dual inhibition of acetylcholinesterase and butyrylcholinesterase may lead to broader efficacy. As acetylcholinesterase activity decreases with disease progression, the acetylcholinesterase-selective agents may lose their effect, while the dual inhibitors may still be effective due to the added inhibition of butyrylcholinesterase. However, this has not been demonstrated clinically. 

Consistent decreases in plasma cholinesterase may not have been observed in rats and dogs because they were treated with lower doses of diisopropyl methylphosphonate. In general, depression of plasma cholinesterase, also known as pseudocholinesterase or butyrylcholinesterase, is considered a marker of exposure rather than an adverse effect. Depression of cholinesterase activity in red blood cells (acetylcholinesterase) is a neurological effect thought to parallel the inhibition of brain acetylcholinesterase activity. It is considered an adverse effect. Acetylcholinesterase is found mainly in nervous tissue and erythrocytes. Diisopropyl methylphosphonate was not found to inhibit RBC... 

Although this study (Hart 1980) did not identify an effect level, the NOAEL is below the LOEL found in all studies examining the toxicity of diisopropyl methylphosphonate. The LOEL for diisopropyl methylphosphonate is 262 mg/kg/day for male mink and 330 mg/kg/day for female mink (Bucci et al. 1997), doses at which statistically significant decreases in plasma cholinesterase (butyrylcholinesterase) but not RBC cholinesterase (acetylcholinesterase) activity were observed (Bucci et al. 1997). In general, a decrease in plasma cholinesterase activity is considered to be a marker of exposure rather than a marker of adverse effect, while a decrease in RBC acetylcholinesterase activity is a neurological effect thought to parallel the inhibition of brain acetylcholinesterase activity and is thus considered an adverse effect. Diisopropyl methylphosphonate was not found to inhibit red blood cell cholinesterase at doses at which plasma cholinesterase was significantly inhibited. No effects were observed in males at 45 mg/kg/day (Bucci et al. 1997) or at 63 mg/kg/day (Bucci et al. 1994), and no effects were observed in females at 82 mg/kg/day (Bucci et al. 1994), or at 57 mg/kg/day (Bucci et al. 1997). 

The inhibition of two cholinesterase activities in blood can also be used to confirm exposure to certain organophosphate ester compounds. Red blood cell acetylcholinesterase is the same cholinesterase found in the gray matter of the central nervous system and motor endplates of sympathetic ganglia. Synonyms for this enzyme include specific cholinesterase, true cholinesterase, and E-type cholinesterase. Plasma cholinesterase is a distinct enzyme found in intestinal mucosa, liver, plasma, and white matter of the central nervous system. Synonyms for this enzyme include nonspecific cholinesterase, pseudocholinesterase, butyrylcholinesterase, and S-type cholinesterase (Evans 1986). Nonspecific cholinesterase is thought to be a very poor indicator of neurotoxic effects. 

Most of the pesticide biosensors are designed based on the inhibitory property of enzymes. AChE and butyrylcholinesterase (BChE) are widely used in the development of pesticide biosensors [17, 18], Inhibition leads to a decrease in activity, which... 

Presently available methods to diagnose and biomonitor exposure to anticholinesterases, e.g., nerve agents, rely mostly on measurement of residual enzyme activity of acetylcholinesterase (AChE) and butyrylcholinesterase (BuChE) in blood. More specific methods involve analysis of the intact poison or its degradation products in blood and/or urine. These approaches have serious drawbacks. Measurement of cholinesterase inhibition in blood does not identify the anticholinesterase and does not provide reliable evidence for exposure at inhibition levels less than 20 %. The intact poison and its degradation products can only be measured shortly after exposure. Moreover, the degradation products of pesticides may enter the body as such upon ingestion of food products containing these products. 

Most insecticides, especially the organophosphate group, cause neurotoxicity as their major mode of action. Assessment of the neurotoxicity includes neurochemical endpoints such as cholinesterase (including acetylcholinesterase, which is the major neurotransmitter in vertebrates such as fish, and other enzymes such as butyrylcholinesterase) inhibition and behavioral endpoints such as swimming speed [79]. Studies done in rats show the neurotoxic action of insecticides such as dimethoate, methyl parathion, dichlorvos, ethyl parathion or propoxur after a prolonged exposure [80,81]. 

Esterases that contribute to human drug metabolism fall into three major classes the cholinesterases (acetylcholinesterase, pseudocholinesterase, butyrylcholinesterase, etc.),... 

Inhibition of the two principal human cholinesterases, acetylcholinesterase and pseudocholinesterase, may not always result in visible neurological effects (Sundlof et al. 1984). Acetylcholinesterase, also referred to as true cholinesterase, red blood cell cholinesterase, or erythrocyte cholinesterase is found in erythrocytes, lymphocytes, and at nerve synapses (Goldfrank et al. 1990). Inhibition of erythrocyte or lymphocyte acetylcholinesterase is theoretically a reflection of the degree of synaptic cholinesterase inhibition in nervous tissue, and therefore a more accurate indicator than pseudocholinesterase activity of inhibited nervous tissue acetylcholinesterase (Fitzgerald and Costa 1993 Sundlof et al. 1984). Pseudocholinesterase (also referred to as cholinesterase, butyrylcholinesterase, serum cholinesterase, or plasma cholinesterase) is found in the plasma, serum, pancreas, brain, and liver and is an indicator of exposure to a cholinesterase inhibitor. 

In contrast to acetylcholinesterase, cholinesterase (acylcholine acyl-hydrolase, butyrylcholinesterase, EC 3.1.1.8) exhibits relatively unspecific esterase activity toward choline esters, with abroad specificity relative to the size of the acyl group. The enzyme is synthesized in the liver and can be found in smooth muscle, adipocytes, and plasma. Its physiological role remains partly obscure, but there is evidence that it is present transiently in the embryonic nervous system, where it is replaced in later stages of development by acetylcholinesterase. It has, therefore, been suggested that cholinesterase functions as an embryonic acetylcholinesterase. 

To help the reader gain a better understanding of the three-dimensional structure of the catalytic site of an esterase, Fig. 3.8 presents the 3D structure of human butyrylcholinesterase (EC 3.1.1.8) obtained by homology modeling [42], The overall structure of the enzyme is shown in Fig. 3.8, a, while Fig. 3.8,b shows a closeup of the active site with the catalytic triad highlighted and the close spatial relationship of the Ser-His-Glu residues revealed. 

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