Purification of cholesterol gallstone inhibitory and promoting glycoproteins in bile and its clinical significance

Holzbach，Groen等人报道，人胆汁中存在促进或抑制胆汁中胆固醇晶体形成的糖蛋白。Holzbach等人报道了一种用分光光度法检测胆固醇成核和生长的新方法，该方法可以半定量地测定这些糖蛋白对胆汁中胆固醇晶体形成的促进或抑制作用。我们通过凝胶层析、亲和层析(ConA和Helix Pomatia Lectin柱)和高效液相色谱柱(离子交换柱)等一系列步骤纯化了糖蛋白。用胆固醇成核和生长实验检测了从人胆汁中纯化的糖蛋白的促进和抑制活性。结果表明，具有促进胆汁胆固醇结晶形成活性的糖蛋白主要存在于ConA凝集素结合蛋白组分中，而具有抑制活性的糖蛋白主要存在于鸡冠花凝集素结合蛋白组分中；对胆汁中胆固醇结晶有促进作用的糖蛋白组分在SDS-PAGE中含有4种不同的蛋白质，其相对分子质量在30~120kD之间。这些糖蛋白正在进一步提纯中。

Presence of glycoproteins in human bile which promote or inhibit cholesterol crystal formation in bile has been reported by Holzbach, Groen et al,. Holzbach et al, has reported a new method of cholesterol nucleation and growth assay using spectrophotometer, which can semi-quantify the promoting or inhibitory effects of those glycoproteins on cholesterol crystal formation in bile. We purified glycoproteins by series of procedures using gel chromatography, affinity chromatography(Con A and Helix pomatia lectin columns), and HPLC (ion exchanger column). Promoting and inhibitory activity of purified glycoproteins from human bile was examined by using cholesterol nucleation and growth assay. As the results, glycoproteins which showed promoting activity on cholesterol crystal formation in bile were mainly found in Con A lectin bounding protein fraction and those showed inhibitory activity were found in Helix pomatia lectin bounding fraction.The glycoprotein fraction which exhibited promoting effect on the cholesterol crystal formation in bile contained four different proteins on SDS-PAGE electrophoresis whose molecular weights ranged from 30 kD to 120 Kd. These glycoproteins are under further purification.