液-液分配色谱法在同时纯化性激素结合球蛋白和皮质类固醇结合球蛋白中的应用。

Application of liquid-liquid partition chromatography in the simultaneous purification of sex-hormone-binding globulin and corticosteroid-binding globulin.

作者信息

Heubner A, Belovsky O, Müller W, Grill H J, Manz B, Juchem M, Pollow K

机构信息

Abteilung für Experimentelle Endokrinologie, Johannes Gutenberg Universität Mainz, F.R.G.

出版信息

J Chromatogr. 1987 Jun 26;397:419-34. doi: 10.1016/s0021-9673(01)85027-5.

DOI:10.1016/s0021-9673(01)85027-5

PMID:2821037

Abstract

Two human serum proteins, corticosteroid-binding globulin (CBG) and sex-hormone-binding globulin (SHBG), were purified to homogeneity by the application of a combination of three different modes of chromatography. Human pregnancy serum was fractionated with ammonium sulphate. SHBG (50% pellet) and CBG (80% pellet) were then purified by affinity chromatography on tresyl-activated Sepharose with 15-aminopentadecanoic acid (for SHBG) and 1,12-diaminododecane (for CBG) as spacers and 17 zeta-aminoethyl-5 alpha-androstan-3 beta,17-diol (for SHBG) and 17 alpha-hydroxy-4-androsten-3-one-17 beta-carboxylic acid (for CBG) as specific ligands for these two proteins. The eluate was injected into a Mono Q anion-exchange column. Fractions containing SHBG or CBG were finally purified by liquid-liquid chromatography on Lipar-Gel 750. This chromatographic sequence clearly separated SHBG and CBG from other proteins, mainly serum albumin, without a loss of protein or binding activity.

摘要

通过应用三种不同色谱模式的组合，将两种人血清蛋白，即皮质类固醇结合球蛋白（CBG）和性激素结合球蛋白（SHBG）纯化至同质。用人妊娠血清进行硫酸铵分级分离。然后，以15 - 氨基十五烷酸（用于SHBG）和1,12 - 二氨基十二烷（用于CBG）作为间隔臂，以17ζ - 氨基乙基 - 5α - 雄甾烷 - 3β,17 - 二醇（用于SHBG）和17α - 羟基 - 4 - 雄烯 - 3 - 酮 - 17β - 羧酸（用于CBG）作为这两种蛋白质的特异性配体，通过在三嗪基活化的琼脂糖凝胶上进行亲和色谱法纯化SHBG（50%沉淀）和CBG（80%沉淀）。将洗脱液注入Mono Q阴离子交换柱。含有SHBG或CBG的级分最后通过在Lipar - Gel 750上进行液 - 液色谱法纯化。该色谱序列清楚地将SHBG和CBG与其他蛋白质（主要是血清白蛋白）分离，且蛋白质或结合活性没有损失。