Acetylcholinesterase, purification

Ciliv, G., and P.T. Oezand. 1972. Human erythrocyte acetylcholinesterase purification, properties and kinetic behavior. Biochim Biophys Acta 284 136. 

Acetylcholinesterase. Purification of acetylcholinesterase has always been problematical with complex separation procedures proving less than satisfactory. A simple one-step purification for acetylcholinesterase from the electric organ of Electrophorus electricus and bovine erythrocyte... 

Ott, R, Membrane acetylcholinesterases Purification, molecular properties and interactions with amphyphilic environments, Biochem. Biophys. Acta, 822, 375, 1985. 

Griffiths, G. and Pritchard, D.I. (1994) Purification and biochemical characterisation of acetylcholinesterase (AChE) from the excretory/secretory products of Trichostrongylus colubriformis. Parasitology 108, 579-586. 

Grigg, M.E., Tang, L., Hussein, A.S. and Selkirk, M.E. (1997) Purification and properties of monomeric (Gl) forms of acetylcholinesterase secreted by Nippostrongylus brasiliensis. Molecular and Biochemical Parasitology 90, 513-524. 

S. Estrada-Mondaca and D. Fournier, Stabilization of recombinant Drosophila acetylcholinesterase, Prot. Exp. Purif., 12 (1998) 166-172. 

Organophosphates are active not only against acetylcholinesterase but also serine proteases - which is obviously due to the shared catalytic mechanism. DFP is actually being used as a protease inhibitor in biotechnology. Another inhibitor that shares its mode of action but is less dangerous (because it is not volatile, and the enzyme adducts it forms are less stable) is PMSF (phenylmethylsulfone fluoride). You may have encountered it in one or the other research lab it is commonly added to crude cell extracts in order to minimize enzymatic breakdown of proteins during purification. 

A similar procedure has been described for the purification of acetylcholinesterase from the electric organ of Torpedo marmorato [68]. The inhibitor, l-(N,N,N-trimethylammonium)-6-hexylamine bromide hydrobromide, was coupled to agarose through a C30 spacer and the resulting affinity media gave an homogenous enzyme fraction with a 93% yield. 

Affinity chromatography is one of the most effective procedures for enzyme purification and yet, with the notable exceptions of Picard (PIO) and of La Du and his co-workers (L2), it has received scant attention for the purification of cholinesterase. Its value in purifying acetylcholinesterase from both fish electric organ and erythrocyte membrane is discussed in an excellent review by Rosenberry (R8). In affinity... 

L28. Leuzinger, W., and Baker, A. L., Acetylcholinesterase. I. Large-scale purification, homogeneity, and amino acid analysis. Proc. Natl. Acad. Sci. U.S.A. 57, 446-451 (1967). 

ColUnson SK, Emody L, Muller KH, Trust TJ, Kay WW (1991) Purification and characterization of thin, aggregative fimbriae from Salmonella enteritidis. J Bacteriol 173 4773 781 Cordes MH, Walsh NP, McKnight CJ, Sauer RT (2003) Solution structure of switch Arc, a mutant with 3(10) helices replacing a wild-type beta-ribbon. J Mol Biol 326 899-909 Cottingham MG, Hollinshead MS, Vaux DJ (2002) Amyloid fibril formation by a synthetic peptide from a region of human acetylcholinesterase that is homologous to the Alzheimer s amyloid-beta peptide. Biochemistry 41 13539-13547... 

Purification of acetylcholinesterase from the head-tissues of house flies Purification of the glycogen-debranching enzyme (amylo-l,6-glucosidase) from do sh muscle... 

Macroporous agarose cross-linked with epichlorohydrin has been reduced and then activated with cyanogen bromide to introduce cyclic imidocarbonate groups this material reacted with the 3- and 4-isomers of A-(6-aminohexanoyl)-1-aminophenyltrimethylammonium bromide hydrobromide to give affinants that were used in the purification of acetylcholinesterase. ... 

Purification of serine esterases and proteases application to eel acetylcholinesterase and human serum plasminogen Purification by immuno HPAC application to human serum albumin... 

Figure 4. Purification of eel acetylcholinesterase on ACA-Separon-immobilized trimethyl (p-aminophenyl) ammonium chloride. HPAC column, ACA-Separon-immobilized TAPA (40 /uiud/g dry gel), 25 cm x 0.8 cm sample applied. 0.5 ml eel extract containing about 35 units enzyme and 9.5 mg protein flow-rate, 1 ml/min starting solvent, 0.01 M sodium phosphate buffer, (pH 7.2) eluant, 1 M NaCI in buffer, (—) protein A2S4, hatched regions indicate enzyme activity. Reproduced from (18) with permission.

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