Solubilization, purification, and characterization of a truncated form of rat hepatic squalene synthetase.
作者： I Shechter , E Klinger , M.L. Rucker , R.G. Engstrom , J.A. Spirito
DOI: 10.1016/S0021-9258(18)42489-1
关键词:
摘要: Rat hepatic microsomal squalene synthetase (EC 2.5.1.21) was induced 25-fold by feeding rats with diet containing the hydroxymethylglutaryl-coenzyme A reductase inhibitor, fluvastatin, and cholestyramine, a bile acid sequestrant. soluble protein an estimated mass of 32-35 kDa, as determined gel filtration chromatography on Sephacryl S-200 column, solubilized out microsomes controlled proteolysis trypsin. Approximately 25% activity recovered in form. The enzyme purified to homogeneity utilizing series column purification steps DEAE-cellulose, hydroxylapatite, phenyl-Sepharose sequentially. showed single band sodium dodecyl sulfate-polyacrylamide electrophoresis. Initial kinetic analysis indicated S0.5 values for trans-farnesyl diphosphate 1.0 microM NADPH 40 microM. Vmax respect calculated at 1.2 mumol/min/mg. NADH also serves substrate reaction value 800 Western blot rabbit antisera raised against purified, trypsin-truncated isolated 32-33 kDa intact about 45-47 kDa.
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