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目的 纯化并比较各种成核效应蛋白的活性,筛选最具病理学意义的促成核因子。方法 通过ConA-Sepharose 4B亲和层析和Sephadex G-200、Sephadex G-100分子筛凝胶过滤以及超速离心法从胆固醇性结石病人胆汁中分离提纯70 kDa、200 kDa促成核蛋白以及33.5 kDa泡蛋白,在模拟胆汁体系中观察成核因子促进胆固醇结晶生长、析出过程,比较其成核活性。结果 四种候选的成核效应蛋白中,33.5 kDa泡蛋白是活性最强的促成核因子,成核活性强弱依次为33.5 kDa泡蛋白>200 kDa糖蛋白≈α1-酸性糖蛋白>70 kDa糖蛋白,当浓度为100／μg／ml时,33.5 kDa泡蛋白成核活性指数Ig、Ic、It分别为1.52、1.63、0.57,并随着蛋白浓度的增加,促成核活性亦随之增强(F=678.18,P<0.05),且呈剂量依赖效应。结论 胆汁中存在多种促成核作用的糖蛋白,其中33.5 kDa泡蛋白可能与胆石形成关系最为密切。
Objective To purify and compare the activity of various nucleating effector proteins and screen for the most pathogenic promoters of nuclear factor. Methods The 70 kDa, 200 kDa prokaryotic protein and 33.5 kDa protein were isolated and purified from the bile of patients with cholesterol gallstone by ConA-Sepharose 4B affinity chromatography, Sephadex G-200, Sephadex G-100 molecular sieve gel filtration and ultracentrifugation , Observed in the simulated bile system nucleating factor to promote the growth of cholesterol crystals, precipitation process, compare its nucleation activity. Results 33.5 kDa vacuolarin was the most active nuclear factor of the four candidate nucleolar effector proteins. The nucleation activity was 33.5 kDa, the order of the protein was> 200 kDa glycoprotein ≈α1-acid glycoprotein> 70 kDa sugar The Ig, Ic and It of 33.5 kDa protein were 1.52, 1.63 and 0.57 respectively at the concentration of 100 / μg / ml. With the increase of protein concentration, the nucleation activity of F = 678.18, P <0.05), and in a dose-dependent manner. Conclusion There are many kinds of glycoproteins in bile that promote nucleation, among which the 33.5 kDa protein may be most closely related to gallstone formation.