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. 1973 Jul;133(3):521–527. doi: 10.1042/bj1330521

Multiple forms of acetylcholinesterase from human erythrocytes

David L Wright 1,*, David T Plummer 1
PMCID: PMC1177730  PMID: 4733238

Abstract

1. Acetylcholinesterase from human erythrocytes was solubilized with Triton X-100 in strong salt solution and partially purified by (NH4)2SO4 fractionation. This preparation showed three main bands of enzyme activity after electrophoresis on polyacrylamide gel and incubation with either α-naphthyl acetate or acetylthiocholine as enzyme substrate. Two of the multiple forms were completely inhibited by 10μm-eserine and one only partially. Treatment with neuraminidase had no effect on the electrophoretic pattern; therefore sialic acid does not appear to determine or affect the ratios of the acetylcholinesterase multiple forms, unlike those of the serum cholinesterase. 2. Chromatography of the preparation on Sephadex G-200 revealed one major peak of enzyme activity and a suggestion of two minor zones of mol.wt. 546000, 184000 and 93000 (i.e. in the proportion 6:2:1). The main peak was almost completely separated from the Triton X-100 and the overall purification was about 600-fold. Further attempts to purify the enzyme by absorption on calcium phosphate gels were unsuccessful. 3. Electrophoresis of the enzyme preparation on a polyacrylamide gradient for 24h revealed three main bands that corresponded to the three values for molecular weights obtained by column chromatography. After 70h of electrophoresis a further three zones of activity developed making six molecular entities, the molecular weights of which were simple multiples of a monomer, thus resembling the cholinesterase found in serum.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. ALLEN J. M., GOCKERMAN J. ELECTROPHORETIC SEPARATION OF MULTIPLE FORMS OF PARTICLE ASSOCIATED ACID PHOSPHATASE. Ann N Y Acad Sci. 1964 Dec 28;121:616–633. doi: 10.1111/j.1749-6632.1964.tb14230.x. [DOI] [PubMed] [Google Scholar]
  2. Andrews P. The gel-filtration behaviour of proteins related to their molecular weights over a wide range. Biochem J. 1965 Sep;96(3):595–606. doi: 10.1042/bj0960595. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. BERNSOHN J., BARRON K. D., HESS A. R. Multiple nature of acetylcholinesterase in nerve tissue. Nature. 1962 Jul 21;195:285–286. doi: 10.1038/195285a0. [DOI] [PubMed] [Google Scholar]
  4. Bellhorn M. B., Blumenfeld O. O., Gallop P. M. Acetylcholinesterase of the human erythrocyte membrane. Biochem Biophys Res Commun. 1970 Apr 24;39(2):267–273. doi: 10.1016/0006-291x(70)90788-6. [DOI] [PubMed] [Google Scholar]
  5. Froede H. C., Wilson I. B. On the subunit structure of acetylcholinesterase. Isr J Med Sci. 1970 Mar-Apr;6(2):179–184. [PubMed] [Google Scholar]
  6. Gaffney P. J., Jr Human serum cholinesterase. I. Partial purification andnature of the heterogeneity of this system. Biochim Biophys Acta. 1970 Jun 23;207(3):465–476. [PubMed] [Google Scholar]
  7. JACOBSEN C. F., LEONIS J., LINDERSTROM-LANG K., OTTESEN M. The pH-stat and its use in biochemistry. Methods Biochem Anal. 1957;4:171–210. doi: 10.1002/9780470110201.ch5. [DOI] [PubMed] [Google Scholar]
  8. Juul P. Human plasma cholinesterase isoenzymes. Clin Chim Acta. 1968 Feb;19(2):205–213. doi: 10.1016/0009-8981(68)90327-6. [DOI] [PubMed] [Google Scholar]
  9. LOWRY O. H., ROSEBROUGH N. J., FARR A. L., RANDALL R. J. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed] [Google Scholar]
  10. LaMotta R. V., Woronick C. L. Molecular heterogeneity of human serum cholinesterase. Clin Chem. 1971 Mar;17(3):135–144. [PubMed] [Google Scholar]
  11. Lamotta R. V., McComb R. B., Noll C. R., Jr, Wetstone H. J., Reinfrank R. F. Multiple forms of serum cholinesterase. Arch Biochem Biophys. 1968 Mar 20;124(1):299–305. doi: 10.1016/0003-9861(68)90331-7. [DOI] [PubMed] [Google Scholar]
  12. Leuzinger W., Goldberg M., Cauvin E. Molecular properties of acetylcholinesterase. J Mol Biol. 1969 Mar 14;40(2):217–225. doi: 10.1016/0022-2836(69)90470-7. [DOI] [PubMed] [Google Scholar]
  13. Lewis P. R., Shute C. C. The distribution of cholinesterase in cholinergic neurons demonstrated with the electron microscope. J Cell Sci. 1966 Sep;1(3):381–390. doi: 10.1242/jcs.1.3.381. [DOI] [PubMed] [Google Scholar]
  14. Margolis J., Kenrick K. G. Polyacrylamide gel electrophoresis in a continuous molecular sieve gradient. Anal Biochem. 1968 Oct 24;25(1):347–362. doi: 10.1016/0003-2697(68)90109-7. [DOI] [PubMed] [Google Scholar]
  15. Maynard E. A. Electrophoretic studies of cholinesterases in brain and muscle of the developing chicken. J Exp Zool. 1966 Apr;161(3):319–335. doi: 10.1002/jez.1401610303. [DOI] [PubMed] [Google Scholar]
  16. Millar David B., Grafius Melba A. The subunit molecular weight of acetylcholinesterase. FEBS Lett. 1970 Dec 23;12(1):61–64. doi: 10.1016/0014-5793(70)80596-8. [DOI] [PubMed] [Google Scholar]
  17. PAUL J., FOTTRELL P. Tissue-specific and species-specific esterases. Biochem J. 1961 Feb;78:418–424. doi: 10.1042/bj0780418. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. WARREN L. The thiobarbituric acid assay of sialic acids. J Biol Chem. 1959 Aug;234(8):1971–1975. [PubMed] [Google Scholar]
  19. Wright D. L., Plummer D. T. Solubilization of acetylcholinesterase from human erythrocytes by Triton X-100 in potassium chloride solution. Biochim Biophys Acta. 1971 Feb 28;261(2):398–401. doi: 10.1016/0304-4165(72)90064-5. [DOI] [PubMed] [Google Scholar]

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