Grant G A, Keefer L M, Bradshaw R A
J Biol Chem. 1978 Apr 25;253(8):2724-6.
A method is described for the preparation of homogeneous D-3-phosphoglycerate dehydrogenase from chicken liver in amounts sufficient for structural studies. The procedure utilizes ammonium sulfate precipitation, blue dextran-Sepharose chromatography, ion exchange chromatography on phosphocellulose, and crystallization. Previous reports of instability of the enzyme have been shown to be due to proteolysis in the crude extract which can be effectively prevented by leupeptin. The purified enzyme is a basic protein with a pI of 8.95 as measured by isoelectric focusing. The extinction coefficient at 278 nm of a 1% solution is 5.3.
本文描述了一种从鸡肝中制备足够量用于结构研究的均一D-3-磷酸甘油酸脱氢酶的方法。该方法利用硫酸铵沉淀、蓝色葡聚糖-琼脂糖凝胶色谱、磷酸纤维素离子交换色谱和结晶。先前关于该酶不稳定的报道已表明是由于粗提取物中的蛋白水解作用，而亮肽素可有效防止这种情况。通过等电聚焦测量，纯化后的酶是一种碱性蛋白，其pI为8.95。1%溶液在278nm处的消光系数为5.3。
