体外培养伊氏锥虫的可变表面糖蛋白的分离与鉴定

ISOLATION AND CHARACTERIZATION OF VARIABLE SURFACE GLYCOPROTEIN OF TRYPANOSOMA EVANSI IN VITRO

体外培养后分离出的3个伊氏锥虫变异群体,经环磷酰胺处理大鼠(CY 大鼠)扩增收虫。虫体纯化后超声波裂解,超速离心制成可溶性抗原,经 Sephadex G-50脱盐和 DE-52离子交换层析提纯可变表面糖蛋白(VSG)。经 SDS-PAGE 鉴定,所提纯的3个 VSG 分子量均为40500。用日立835-50型高速氨基酸组成分析仪分析,3个 VSG 均由 Asp,Pro,Thr,Ser,Glu,Gly,Ala,Cys,Val,Met,Ile,Leu,Tyr,Phe,Lys,His,Arg共17种氨基酸组成,但在组成比例上差异明显,尤其表现在 Pro,Met,Leu,Ile.Tyr 和 Clu。经 Western blot-ting 鉴定,提纯的3种 VSG 是3种不同的特异性抗原。

Three VAT-derived populations were expanded in cyclophosphamide treated(CY)rats for experimen- tal use.Purified trypanosomes were broken by high-speed homogenization and centrifugated by high g to form soluble proteins.The soluble proteins were desalted on a column of Sephadex G-50 prior to chromatography on a DE-52 column.The material in the first peak constituted the purified VSG.The molecular weight of the purified VSG was 40 500 by detection of SDS-PAGE.The differences of three VSGs in molecular weight were small.Three VSGs were composed of 17 amino acids by analysis of Model 835-50 high speed amino acid analyzer.They were Asp,Pro,Thr, Ser,Glu,Gly,Ala,Cys,Val,Met,He,Leu,Tyr,Phe,Lys,His,and Arg.The differences in the proportion of amino acid were most striking especially in Pro,Mei,Leu,He,Tyr,and Glu.Three purified VSGs were identified as specific antigens of three variant coats by Western blotting.