Reconstitution of human erythrocyte membrane acetylcholinesterase in phospholipid vesicles. Analysis of the molecular forms by cross-linking studies.

Unilamellar lipid vesicles were formed upon removal of Triton X-100 with Amberlite XAD-2 from a mixture of egg phosphatidylcholine and Triton-solubilized pure human erythrocyte membrane acetylcholinesterase. A majority of large (230 nm diameter) vesicles together with a minor population of smaller (30 nm diameter) strictures were observed in freeze-fracture electron micrographs. Reconstitution experiments performed with [phenyl-3H(n)]-Triton X-100 showed that only one detergent molecule per 600 lipid molecules was present in the vesicles. Density gradient centrifugation showed co-sedimentation of acetylcholinesterase with the lipid vesicles. About 60% of the incorporated enzyme was directed towards the vesicle exterior and could be partially degraded by papain. Mainly dimeric acetylcholinesterase was found when the reconstituted or, alternatively, the lipid-free but Triton-solubilized enzyme were cross-linked with glutaraldehyde. Aggregates were observed when the detergent-depleted oligomeric forms of the enzyme were cross-linked. The results thus indicate that mainly the dimeric enzyme form is present in a phospholipid environment.