Muensch H, Goedde H W, Yoshida A
Eur J Biochem. 1976 Nov 1;70(1):217-23. doi: 10.1111/j.1432-1033.1976.tb10972.x.
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.
通过两步法对人血清胆碱酯酶的主要C4成分进行了高度纯化，该两步法包括在DEAE - 纤维素上进行色谱分离和制备性圆盘电泳。最终产物纯化了约8000倍，产率为64%。通过8M尿素聚丙烯酰胺圆盘电泳和在5M盐酸胍中的沉降平衡离心法确定了亚基结构。发现C4酶具有四聚体结构。这些亚基在大小和电荷上相等，分子量与天然血清中的C1酶相当。对主要的C4酶和次要的C1酶进行了“活性酶离心”。发现在底物存在下，C4酶是四聚体，C1酶是单体。从结合的二异丙基磷酸与蛋白质的摩尔比测量二异丙基磷酰氟结合位点的数量。发现C4酶的结合位点值接近两个。
