Purification of a presynaptic membrane protein that mediates a calcium-dependent translocation of acetylcholine.

A protein, which we call "mediatophore," that mediates calcium-dependent release of acetylcholine from proteoliposomes has been purified from the presynaptic plasma membrane. About 250 micrograms of this material was obtained from 500 g of Torpedo marmorata electric organ. Precipitation of the protein and subsequent removal of associated lipids inactivated the protein, which then became water soluble; this permitted evaluation of its Stokes radius (52 A) and its sedimentation coefficient (9.8 +/- 0.75 S) and, hence, an approximate molecular mass of 210 +/- 16 kDa could be determined. PAGE analysis showed that the protein is made of 17-kDa subunits, not linked by disulfide bonds. When this material was observed by electron microscopy after negative staining, the apparently pentameric structures had an average diameter of about 7 nm.