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The large-scale isolation of an acetylcholinesterase from the haemolymph of the sea mussel mytilus edulis by means of fractional (NH4)2SO4 precipitation, affinity chromatography and isoelectric focusing is described. The yield is about 20% of the initial enzyme activity with a specific activity of 1727 nkat/mg protein. Purification steps are followed by disc electrophoresis, analytical isoelectric focusing and immunoelectrophoresis, showing the enzyme to be homogeneous with an isoelectric point of 4.81 +/- 0.02 and to have an apparent molecular weight of 245,000, which is composed by six equal subunits as judged by C- and N-terminal amino acid analysis. The acetylcholinesterase is a metalloprotein containing three Fe2 ions per subunit (molecular weight 40625); this consists of 351 amino acid and six sugar residues, but contains no sialic acid. As calculated from CD data, the enzyme has about 55% helical structure.

By use of different inhibitors as well as atomic absorption spectrophotometry it has been shown that the haemolymph-acetylcholinesterase (E. c. 3.1.1.7) of the sea mussel Mytilus edulis is a metalloprotein containing 2,95 Fe2+-ions per subunit. All inhibitors used (1,10-phenanthroline, salicylic aldehyde, 2,2'-dipyridyl, 8-hydroxyquinoline) showed a non-competitive inhibition, which was not pH-dependent. Some divalent cations caused a marked increase of the enzyme activity, some heavy metals inhibited the enzyme almost completely; monovalent inorganic cations did not influence the enzyme at all. Besides NaF and Na2SiF6, which showed a non-competitive inhibition comparable to the inhibition observed with the chelating agents, and NaN3, whose mode of action was not identifiable, no inhibition by different mono- and divalent inorganic anions was to be observed. Ammonium ions caused no enzyme inhibition, but length the inhibition power of substituted ammonium ions increased with an increasing C-chain. The influence of some organic solvents on the enzyme activity is demonstrated.

A cholinesterase deriving from the hemolymph of the mussel Mytilus edulis was partially purified by use of gel-permeation and ion-exchange chromatography; the specifity to different substrates or inhibitors indicates clearly the occurrence of a "true" acetylcholinesterase.