Acetylcholinesterase variants

Deutsch, V.R., Pick, M., Perry, C., Grisaru, D., Hemo, Y., Golan-Hadari, D., Grant, A., Eldor, A., Soreq, H. (2002). The stress-associated acetylcholinesterase variant AChE-R is expressed in human CD34(+) hematopoietic progenitors and its C-terminal peptide ARP promotes their proliferation. Exp. Hematol. 30 1153-61. 

Grisara, D., Deutsch, V., Shapira, M., Pick, M., Stemfeld, M., Melamed-Book, N., Kaufer, D., Galyam, N., Gait, M.J., Owen, D., Lessing, J.B., Eldor, A., Soreq, H. (2001). ARP, a peptide derived from the stress-associated acetylcholinesterase variant, has hematopoietic growth promoting activities. Mol. Med. 7 93-105. 

Another biosensor was developed for AN-a(s) by Devic et al. (2002) that addressed the lack of sensitivity and specificity of detection of some bioassays. An acetylcholinesterase was engineered to increase its sensitivity and a combination of mutants was used to obtain increased analyte specificity. The sensitivity of detection achieved was reported to be below 1 nM/L. By using a four-mutant set of acetylcholinesterase variants, two of which are sensitive to AN-a(s) and two are sensitive to the insecticides, an improvement in the specificity of the biosensor for the cyanobacterial neurotoxin was achieved (Devic et al., 2002). 

Other metabolic enzymes that show polymorphic differences in that they can occur as genetic high-activity and low-activity variants include acetylcholinesterase, butyrylcholinesterases, flavin-dependent monooxygenase, alcohol dehydrogenase, epoxide hydrolase, and arylesterase (Beltoft et al. 2001). 

Modulated splicing-associated gene expression in PI9 cells expressing distinct acetylcholinesterase splice variants. J. Neurochem. 97 (Suppl. 1) 24-34. 

Dori, A., Cohen, J., Silverman, W.F., Pollack, Y., Soreq, H. (2005). Functional manipulations of acetylcholinesterase splice variants highlight alternative splicing contributions to murine neocortical development. Cereb. Cortex 15 419-30. 

Gilboa-Geffen, A., Lacoste, P.P., Soreq, L., Cizeron-Clairac, G., Le Panse, R., Truffault, F., Shaked, I., Soreq, H., Berrih-Aknin, S. (2007). The thymic theme of acetylcholinesterase splice variants in myasthenia gravis. Blood 109 4383-91. 

Perrier, N.A., Salani, M., Falasca, C., Bon, S., Augusti-Tocco, G., Massoulie, J. (2005). The readthrough variant of acetylcholinesterase remains very minor after heat shock, organophosphate inhibition and stress, in cell culture and in vivo. J. Neurochem. 94 629-38. 

Stemfeld, M., Shoham, S., Klein, O., Flores-Flores, C., Evron, T., Idelson, G.H., Kitsberg, D., Patrick, J.W., Soreq, H. (2000). Excess read-through acetylcholinesterase attenuates but the synaptic variant intensifies neurodeterioration correlates. Proc. Natl Acad. Set USA 97 8647-52. 

There is a vast and continually growing literature on these two enzymes, but many papers have not distinguished clearly between them. The purpose of the present survey is to focus attention on human serum cholinesterase and its variants in the light of recent research, and to offer a critical assessment of reports of its physical and chemical properties and clinical and toxicological applications. The methodologies of cholinesterase assays and the older literature on the variant enzymes will not be considered in detail Relevant references may be found in several authoritative reviews (B15, D5, F4, G13, G16, H5, K5, LIO, L24, Ul, W35). Acetylcholinesterase, also, has been extensively reviewed (Nl, S22, S23, W29) and it will be discussed here only in order to make particular points of comparison with cholinesterase. 

A procedure for identifying certain cholinesterase variants was proposed by Dietz et al. (D15). After a period during which comments and discussion were offered by others working in the field, the method was published in Selected Methods of Clinical Chemistry (D16). This method is based upon the Ellman reaction (ElO), which was used by Ellman et al. (Ell) for the assay of acetylcholinesterase, and by Garry and Routh (G9) for the assay of serum cholinesterase. In these assay procedures, a thiocholine ester is used as the substrate. The thiocholine produced upon hydrolysis reacts with 5,5 -dithiobis(2-nitrobenzoic acid) (DTNB) to yield 5-thio-2-nitrobenzoate anion and other products. The rate of the reaction may be determined by measuring the rate at which... 

Figure 9.2 Devonshire and Sawicki (1979) at Rothamstead Experimental Station, Hertfordshire, U.K., found seven variants of the aphid Myzus persicae with different resistances to parathion. Excess production of a carboxylesterase as a result of one or several gene duplications was found to be the resistance mechanism. High levels of carboxylesterases take paraoxon away from acetylcholinesterase so that aphids become resistant.

Many individuals have genetic susceptibility to certain chemicals (Calabrese 1978). The influence of these genetic differences likely produces sub- and supersensitivity to OP insecticides and warfare agents (Russell and Overstreet 1987). Several enzymes with variations or polymorphisms control sensitivity to OPs red blood cell acetylcholinesterase, serum cholinesterase or pseudocholinesterase, lymphocyte neuropathy target esterase or platelet neuropathy target esterase (NTE), serum paroxonase, butyrylcholinesterase, and serum arylesterase (Costa et al. 1999 LaDu 1988 Li et al. 1993 Mutch et al. 1992). Inhibition of red blood cell acetylcholinesterase, in both the central and the peripheral nervous systems, produces acute symptoms (Mutch et al. 1992). Paroxonase and arylesterase further modify the response (LaDu 1988 Li et al. 1993). Variant, inactive butyrylcho-linesterases increase sensitivity to OPs (Lockridge and Masson 2000 Schwarz et al. 1995). OP-induced delayed polyneuropathy results... 

Pharmacokinetics Succinylcholine is composed of two acetylcholine molecules linked end to end. Succinylcholine is metabolized by plasma cholinesterase (butyrylcholinesterase or pseudocholinesterase), which determines the amount of drug reaching the end plate. It has a duration of action of only a few minutes if given as a single dose. Blockade may be prolonged in patients with genetic variants of plasma cholinesterase that metabolize succinylcholine very slowly. Succinylcholine is not rapidly hydrolyzed by acetylcholinesterase. 

Gilboa-Geffen, A., Lacoste, P.P., et al., 2007. The thymic theme of acetylcholinesterase splice variants in myasthenia gravis. Blood 109,4383-4391. 

The activities of two enzymes have been used as biomarkers of effects for OPs, namely acetylcholinesterase (EC 3.1.1.7) and butyrylcholinesterase, sometimes known as pseudocholinesterase (EC 3.1.1.8). The structure and function of these enzymes has been reviewed. " In humans the former is present in red blood cells and the latter in plasma, but such distribution is not true of all species. In dogs, both enzymes are present in plasma with a ratio of butyrylcholinesterase to acetylcholinesterase of 7 1, while in the rat, plasma cholinesterase activity comprises more acetylcholinesterase with a butyrylcholinesterase to acetylcholinesterase activity of 1 3 in males and 2 1 in females in neither blood compartment are the functions of the enzymes fully understood.Because of the possibility of confusion, the terms plasma cholinesterase and erythrocyte cholinesterase as synonyms for butyrylcholinesterase and acetylcholinesterase are to be deprecated, especially when used of enzymes in animals where serious confusion may result. It is often necessary to look in detail at animal studies to see what activity has been measured in each matrix. In particular, it is necessary to look at the substrate(s) used in the assay together with any inhibitors used. Methods for measuring acetylcholinesterase have been reviewed and acetylcholinesterase and butyrylcholinesterase activities can be measured separately. In almost all cases it is the enzyme activity, rather than protein concentration, that is measured and many of the procedures used are variants of the Ellman method. Correct storage of blood samples is important as reactivation of inhibited enzymes ex vivo can occur. 

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