Ito Y, Seno N, Matsumoto I
J Biochem. 1985 Jun;97(6):1689-94. doi: 10.1093/oxfordjournals.jbchem.a135227.
New procedures to immobilize high concentrations of protein ligands by reductive amination on two types of formyl-carriers (I & II) having different spacer lengths were investigated in order to prepare stable and high-capacity adsorbents essential for efficient affinity chromatography. Formyl-carrier (I) was prepared by reductive amination with glutaraldehyde of the amino-carrier obtained on amination of an epoxy-activated carrier. Formyl-carrier (II) was prepared by sodium metaperiodate (NaIO4) treatment of a glyceryl-carrier obtained on hydrolysis of an epoxy-activated carrier. Especially high concentrations of protein ligands were immobilized on formyl-Sepharose 4B (I) under very mild conditions (pH 7.0, 4 degrees C). A series of lectins, one of the most useful classes of group-specific ligands, was successfully immobilized by the procedures. Concanavalin A-Sepharose 4B (I) thus obtained exhibited an adsorption capacity five times greater than that of concanavalin A-Sepharose 4B made by Pharmacia Fine Chemicals, and could be repeatedly used over twenty times without a significant reduction in its adsorption capacity.
研究了通过还原胺化作用将高浓度蛋白质配体固定在两种具有不同间隔长度的甲酰基载体（I和II）上的新方法，以制备高效亲和色谱所需的稳定且高容量的吸附剂。甲酰基载体（I）是通过对环氧活化载体胺化后得到的氨基载体与戊二醛进行还原胺化制备的。甲酰基载体（II）是通过对环氧活化载体水解后得到的甘油基载体用偏高碘酸钠（NaIO4）处理制备的。在非常温和的条件下（pH 7.0，4℃），尤其能将高浓度的蛋白质配体固定在甲酰化琼脂糖4B（I）上。通过这些方法成功固定了一系列凝集素，凝集素是最有用的一类组特异性配体。由此得到的伴刀豆球蛋白A-琼脂糖4B（I）的吸附容量比Pharmacia Fine Chemicals生产的伴刀豆球蛋白A-琼脂糖4B高五倍，并且可以重复使用二十多次而其吸附容量没有明显降低。
