Document Detail

Membrane-bound choline acetyltransferase of the torpedo has characteristics of an integral membrane protein and can be solubilized by proteolysis.
MedLine Citation:
PMID:  1504766     Owner:  NLM     Status:  MEDLINE    
Due to Triton X-114 fractionation of synaptosomes isolated from the electric organ of the fish Torpedo, the existence of a hydrophilic and an amphiphilic form of the enzyme choline-O-acetyltransferase (ChAT) was revealed. Amphiphilic ChAT which represents about 10% of total enzyme activity in synaptosomes, reached 40% of ChAT activity measured in preparations of synaptosomal plasma membranes (SPM) which were washed with solutions of increasing ionic strength. ChAT activity bound to washed SPM could be partially solubilized using proteinase K but not phospholipase C. No ChAT solubilization occurred by treating intact synaptosomes with proteinase K. Water/Triton X-114 partition coefficients of hydrophilic and amphiphilic ChAT were found to be 6.5 and 0.17, respectively. Sedimentation coefficients determined by centrifugation in linear density gradients of sucrose containing Triton X-100, were 4.2S and 4.4S for amphiphilic and hydrophilic ChAT, respectively. On the other hand, removal of Triton X-114 from the detergent phase containing amphiphilic ChAT activity led to enzyme aggregation. Finally, amphiphilic ChAT was slightly more acidic (pH 6.6) than was hydrophilic enzyme (6.8-7.0). We conclude that in Torpedo synaptosomes two forms of ChAT activity, a soluble and a membrane-bound form, are indeed present which differ in their hydrophobicity. The soluble form is hydrophilic. The membrane-bound form is amphiphilic and it aggregates upon removal of detergent. These are two characteristics of integral membrane proteins. Membrane-bound ChAT is most probably intracellularly oriented and not bound to membrane through a 'receptor' protein.
L Eder-Colli; P A Briand; Y Dunant
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Publication Detail:
Type:  Journal Article; Research Support, Non-U.S. Gov't    
Journal Detail:
Title:  Brain research     Volume:  573     ISSN:  0006-8993     ISO Abbreviation:  Brain Res.     Publication Date:  1992 Feb 
Date Detail:
Created Date:  1992-09-18     Completed Date:  1992-09-18     Revised Date:  2006-11-15    
Medline Journal Info:
Nlm Unique ID:  0045503     Medline TA:  Brain Res     Country:  NETHERLANDS    
Other Details:
Languages:  eng     Pagination:  284-92     Citation Subset:  IM    
Department of Pharmacology, Centre Médical Universitaire, Geneva, Switzerland.
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MeSH Terms
Acetylcholinesterase / metabolism
Centrifugation, Density Gradient
Choline O-Acetyltransferase / isolation & purification,  metabolism*
Electric Organ / enzymology*
Horseradish Peroxidase / metabolism
Intracellular Membranes / enzymology*
Isoenzymes / isolation & purification,  metabolism*
L-Lactate Dehydrogenase / metabolism
Membrane Proteins / isolation & purification,  metabolism*
Polyethylene Glycols
Synaptosomes / enzymology*
Reg. No./Substance:
0/Detergents; 0/Isoenzymes; 0/Membrane Proteins; 0/Polyethylene Glycols; 9036-19-5/Nonidet P-40; EC Dehydrogenase; EC 1.11.1.-/Horseradish Peroxidase; EC O-Acetyltransferase; EC 3.-/Hydrolases; EC

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