Human-serum cholinesterase subunits and number of active sites of the major component.

Abstract

The major C4 component of human serum cholinesterase was highly purified by a two-step procedure involving chromatography on DEAE-cellulose and preparative disc electrophoresis. The final product was about 8 000-fold purified with a yield of 64%. The subunit structure was determined by 8M urea polyacrylamide disc electrophoresis and by the sedimentation equilibrium centrifugation method in 5M guanidine hydrochloride. It was found that the C4 enzyme has a tetrameric structure. The subunits are equal in size and charge and a molecular weight comparable to that of the C1 enzyme from native serum. The major C4 enzyme and the minor C1 enzyme were subjected to an 'active enzyme centrifugation'. It was found that the C4 enzyme was a tetramer and the C1 enzyme was a monomer in the presence of substrate. The number of diisopropylphosphofluoridate-binding sites was measured from the molar ratio of bound diisopropylphosphate to protein. A value close to two binding sites was found for the C4 enzyme.