Ghosh S, Grogan W M
Department of Biochemistry and Molecular Biophysics, Medical College of Virginia, Virginia Commonwealth University, Richmond 23298-0614.
Lipids. 1991 Oct;26(10):793-8. doi: 10.1007/BF02536160.
A rat liver cytosolic cholesteryl ester hydrolase (CEH) was purified 12,600-fold by ammonium sulfate precipitation, cation exchange chromatography and gel permeation high-performance liquid chromatography, with an overall yield of 20%. Its properties are compared to those of pancreatic CEH, with which it has sometimes been identified. Liver CEH exhibited a single silver stained band following SDS-polyacrylamide gel electrophoresis (Mr = 66 kDa), was activated by 0.5-10 mM taurocholate but was strongly inhibited by higher levels of taurocholate, which activate pancreatic CEH. Whereas bile salts are known to induce formation of a hexamer of pancreatic CEH, in the current study, 0.5 mM taurocholate dissociated a multimeric form of liver CEH to monomer. Liver CEH did not coelute with pancreatic CEH from cation exchange and chromatofocusing columns, exhibited no immunoreactivity with anti-rat pancreatic CEH IgG in Western blots, was not inhibited by anti-rat pancreatic CEH IgG and had a different amino acid composition from pancreatic CEH. In contrast to liver CEH, which is known to be activated by protein kinases A and C, pancreatic CEH was unaffected by cofactors for protein kinase A and was inhibited by cofactors for protein kinase C.
通过硫酸铵沉淀、阳离子交换色谱和凝胶渗透高效液相色谱法，对大鼠肝脏胞质胆固醇酯水解酶（CEH）进行了纯化，纯化倍数达12,600倍，总产率为20%。将其性质与有时被认为与之相同的胰腺CEH的性质进行了比较。肝脏CEH在SDS-聚丙烯酰胺凝胶电泳后呈现一条银染带（Mr = 66 kDa），被0.5 - 10 mM牛磺胆酸盐激活，但被更高水平的牛磺胆酸盐强烈抑制，而更高水平的牛磺胆酸盐会激活胰腺CEH。已知胆汁盐会诱导胰腺CEH形成六聚体，而在本研究中，0.5 mM牛磺胆酸盐使肝脏CEH的多聚体形式解离为单体。肝脏CEH与胰腺CEH在阳离子交换柱和色谱聚焦柱上不会共同洗脱，在Western印迹中与抗大鼠胰腺CEH IgG无免疫反应性，不受抗大鼠胰腺CEH IgG抑制，且氨基酸组成与胰腺CEH不同。与已知被蛋白激酶A和C激活的肝脏CEH不同，胰腺CEH不受蛋白激酶A的辅因子影响，且被蛋白激酶C的辅因子抑制。
