草鱼脂多糖结合蛋白的分离纯化(英文)  

Isolation and Purification of Lipopolysaccharide-binding Protein (LBP) from Grass Carp,Ctenopharyngodon idellus

作　　者：黄腾[1] 苏建国[1] 董捷[1] 

机构地区：[1]西北农林科技大学动物科技学院陕西省农业分子生物学重点实验室,陕西杨凌712100

出　　处：《西北农业学报》2010年第1期6-11,共6页Acta Agriculturae Boreali-occidentalis Sinica

基　　金：National Natural Science Foundation of China (30871917);Program for New Century Excellent Talentsin University (NCET-08-0466);the Project-sponsored by SRF for ROCS,SEM(14110104)

摘　　要：脂多糖结合蛋白(lipopolysaccharide binding proteins ,LBP)是机体对革兰氏阴性菌感染产生的可溶性急性期蛋白,结合并提呈LPS给细胞表面的模式识别受体,激发免疫反应。在本试验中,用低浓度嗜水气单胞菌( Aeromonas hydrophila)感染草鱼(Ctenopharyngodon idellus)24 h后,抽取全血,离心,获得血浆。结合LBP活性检测,经硫酸铵沉淀、CMSephadex C-50阳离子交换层析和DEAE Sephadex A-25阴离子交换层析后,分离纯化得到LBP。该蛋白对异硫氰酸荧光素标记的脂多糖(Fluorescein isothiocyanate labeled li-popolysaccharide ,FITC-LPS)有较强的结合能力。在SDS-PAGE电泳后,考马斯亮蓝染色可见3条明显的带,分子量大约为68、53和48 kDa。同时探索一条简便分离纯化脂多糖结合蛋白的方法,为进一步研究LBP的功能奠定了基础。Lipopolysaccharide binding protein （LBP） is a soluble acute phase protein that binds to bacterial LPS to elicit immune responses by presenting the LPS to important cell surface pattern recognition receptors,involved in the acute-phase immunologic response to gram-negative bacterial infections.In present study,fresh whole blood was collected from grass carp （Ctenopharyngodon idellus） challenged with Aeromonas hydrophila for 24h.The plasma was obtained by centrifuge.Integrated activity test of LBP and ammonium sulphate precipitation,CM Sephadex C-50 cation-exchange chromatography and DEAE Sephadex A-25 anion-exchange chromatography,lipopolysaccharide binding proteins （LBPs） were isolated and purified.The results indicated that the purified LBPs had relatively high affinity to Fluorescein isothiocyanate labeled lipopolysaccharide （FITC-LPS）.SDS-PAGE stained with Coomassie Brilliant Blue,visualized that there were three distinct bands with molecular weight of 68kDa,53 kDa and 48 kDa respectively.This research described a simple and maneuverable method for isolation and purification of LBPs and served the further study of LBP functions.

关 键 词：脂多糖结合蛋白 草鱼 分离 纯化 

分 类 号：S917.4[农业科学—水产科学]