Brown R E, Jarvis K L, Hyland K J
Hormel Institute, University of Minnesota, Austin 55912.
Biochim Biophys Acta. 1990 May 1;1044(1):77-83. doi: 10.1016/0005-2760(90)90221-i.
Bovine brain contains a lipid transfer protein that is specific for neutral glycosphingolipids and gangliosides but does not stimulate phospholipid or neutral lipid intermembrane transfer (Brown, R.E., Stephenson, F.A., Markello, T., Barenholz, Y. and Thompson, T.E. (1985) Chem. Phys. Lipids 38, 79-93). This report describes a new procedure for purifying glycolipid transfer protein from bovine brain as well as a characterization of the resulting protein. Chief among the newly introduced approaches are dye-ligand and fast protein cation-exchange liquid chromatography. Other modifications include increasing the overall scale of purification, incorporating a pH precipitation step and adding different proteinase inhibitors. The resulting procedure simplifies and accelerates the purification process while yielding a homogeneous protein. The purified protein has a molecular weight near 23 kDa as estimated by sodium dodecyl sulfate polyacrylamide gel electrophoresis. Chromatofocusing reveals that glycolipid transfer protein activity co-elutes with the 23 kDa protein and has an isoelectric point near pH 9.0. A similar isoelectric point is observed using denaturing isoelectric focusing conditions. The protein's amino acid composition reveals high levels of amino acids with non-polar side chains (48%). Based on the findings reported here and on previously published data, bovine brain glycolipid transfer protein has been compared to other lipid transfer proteins as well as lysosomal sphingolipid activator proteins.
牛脑中含有一种脂质转移蛋白，它对中性糖鞘脂和神经节苷脂具有特异性，但不刺激磷脂或中性脂质的膜间转移（Brown, R.E., Stephenson, F.A., Markello, T., Barenholz, Y. 和 Thompson, T.E. (1985) 《化学与物理脂质》38, 79 - 93）。本报告描述了一种从牛脑中纯化糖脂转移蛋白的新方法以及对所得蛋白质的特性表征。新引入的方法中主要有染料配体和快速蛋白质阳离子交换液相色谱法。其他改进包括扩大纯化的总体规模、加入pH沉淀步骤以及添加不同的蛋白酶抑制剂。所得方法简化并加速了纯化过程，同时得到一种均一的蛋白质。通过十二烷基硫酸钠聚丙烯酰胺凝胶电泳估计，纯化后的蛋白质分子量接近23 kDa。色谱聚焦显示糖脂转移蛋白活性与23 kDa蛋白质共洗脱，其等电点接近pH 9.0。在变性等电聚焦条件下观察到类似的等电点。该蛋白质的氨基酸组成显示出高含量具有非极性侧链的氨基酸（48%）。基于此处报告的发现以及先前发表的数据，已将牛脑糖脂转移蛋白与其他脂质转移蛋白以及溶酶体鞘脂激活蛋白进行了比较。
