凝集素染色分析胆汁成核活性蛋白的研究  

Analysis of biliary pro-nucleating vesicular protein with lectin coupled to a peroxidase

作　　者：项建斌[1] 蔡端[1] 张延龄[1] 马保金[1] 查锡良[2] 王丽影[2] 

机构地区：[1]复旦大学华山医院外科,200040 [2]复旦大学基础医学院生化教研室

出　　处：《中华消化杂志》2002年第3期148-149,共2页Chinese Journal of Digestion

基　　金：国家自然科学基金资助项目 ( 395 70 6 95 )

摘　　要：目的　探讨促成核蛋白中糖链的结构与类型及其在胆石形成中的作用。方法　凝集素探针介导的点印迹法分析相对分子质量为 335 0 0的泡蛋白糖链组成与结构、筛选出最强结合凝集素 ,并借助透射电镜观察泡蛋白参与模拟胆汁泡聚集、融合的微观过程。通过结晶生长试验检测 335 0 0泡蛋白及其脱糖基产物的促成核活性。结果　 335 0 0泡蛋白糖链为DSA强结合、ConA(- ) ,属复杂型多天线聚糖。HRP DSA标记泡蛋白糖链可直接反映泡蛋白促胆汁泡聚集、融合及胆固醇单水结晶析出的动态过程。 335 0 0泡蛋白具有明显的促成核活性 ,其成核时间指数、结晶生长速度指数及结晶最终浓度指数分别为 0 .5 7,1.5 2及 1.6 3(P <0 .0 5 ) ,脱糖基后其成核活性几乎完全丧失。结论　糖链影响 335 0 0泡蛋白促成核活性 ,提示其可能参与成核效应蛋白的致石作用过程。Objective To explore the structure and type of sugar chain in 33 500 pro-nucleating protein, and its role in gallstone formation. Methods The 33 500 vesicular protein was examined by dot-immunobinding assay of lectin coupled to a peroxidase (HRP-DSA,HRP-ConA,HRP-WGA). The morphology of biliary vesicles was observed under transmission electron microscopy in process of massive vesicular aggregation, culmination and crystal formation. The protein and its enzymatic deglycosylation fractions nucleation promoting activity were detected by cholesterol crystal growth assay. Results 33 500 vesicular protein with multiantennary and complicated glycan displayed apparent potency of nucleation promotion, which clearly reflected by HRP-DSA immunobinding, and derived crystal growth curve indices. It, Ig, Ic were presented as 0.57, 1.52, 1.63 respectively, but after treated by N-glycanase enzyme, no promoting activity was found. Conclusions Our data suggest the sugar chain play an important role in pro-nucleating process, and may be involved in the gallstone formation.

关 键 词：糖链 成核效应蛋白 凝集素 胆结石 结石形成 

分 类 号：R657.4[医药卫生—外科学]