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Acetylcholinesterase in Mouse Neuroblastoma NB 2 A Cells: Analysis of Production, Secretion, and Molecular Forms
Author(s) -
Stieger S.,
Bütikofer P.,
Wiesmann U. N.,
Brodbeck U.
Publication year - 1989
Publication title -
journal of neurochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.75
H-Index - 229
eISSN - 1471-4159
pISSN - 0022-3042
DOI - 10.1111/j.1471-4159.1989.tb01865.x
Subject(s) - acetylcholinesterase , biochemistry , chemistry , gel electrophoresis , aché , polyacrylamide gel electrophoresis , enzyme , tetramer , sodium dodecyl sulfate , secretion , microbiology and biotechnology , biology
The mouse neuroblastoma cell line NB 2 A produces cellular and secreted acetylcholinesterase (AChE). After incubation of the cells for 4 days the ratio between AChE secreted into the medium and AChE in the cells was 1:1. The cell‐associated enzyme could be subdivided into soluble AChE (25%) and detergent‐soluble AChE (75%). Both extracts contained predominantly monomelic AChE (4.6S) and minor amounts of tetrameric AChE (10.6S), whereas the secreted AChE in the culture supernatant contained only the tetrameric form. All forms were partially purified by affinity chromatography. It could be demonstrated that the secretary and the intracellular soluble tetramers were hydrophilic, whereas the detergent‐soluble tetramer was an amphiphilic protein. On the other hand the soluble and the detergent‐soluble monomelic forms were amphiphilic and their activity depended on the presence of detergent. By digestion with proteinase K amphiphilic monomelic and tetrameric AChE could be converted to a hydrophilic form that no longer required detergent for catalytic activity. Sodium dodecyl sulfate‐polyacrylamide gel electrophoresis of [ 3 H]diisopropylfluorophosphate‐labelled AChE gave one band at 64 kilodaltons (kD) under reducing conditions and two additional bands at 120 kD and 140 kD under nonreducing conditions.