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来自胃肠道线虫寄生虫捻转血矛线虫肠道的一种β-半乳糖苷结合蛋白(半乳糖凝集素)的克隆与特性分析

Cloning and characterization of a beta-galactoside-binding protein (galectin) from the gut of the gastrointestinal nematode parasite Haemonchus contortus.

作者信息

Newlands G F, Skuce P J, Knox D P, Smith S K, Smith W D

机构信息

Moredun Research Institute, International Research Centre, Penicuik, UK.

出版信息

Parasitology. 1999 Nov;119 ( Pt 5):483-90. doi: 10.1017/s003118209900503x.

Abstract

A cDNA encoding a beta-galactoside-binding lectin (galectin) was identified by immunoscreening a Haemonchus contortus cDNA library with antisera from lambs vaccinated with a membrane protein complex (H-gal-GP) derived from the parasites' gut. The cDNA sequence, exhibiting a tandem repeat structure and designated Hco-gal-2, showed significant levels of similarity with galectins from several species of nematode as well as mammalian galectin type 4. Native galectin was preferentially extracted from the H-gal-GP complex and also from an insoluble membrane fraction prepared from adult worms using lactose-agarose affinity chromatography. The affinity-purified material had apparent molecular mass of around 35 kDa with 3 distinct bands visible on SDS-PAGE. All 3 bands were identified as galectins by reaction with antiserum raised to recombinant Hco-GAL-2 on Western blot. To determine whether H. contortus galectins have any protective capacity against infection lambs were vaccinated with affinity-purified galectin and subsequently given a single challenge infection. Vaccination did not confer any protection against infection with H. contortus as judged by faecal egg output or worm counts.

摘要

通过用来自接种了源自寄生虫肠道的膜蛋白复合物(H-gal-GP)的羔羊的抗血清对捻转血矛线虫cDNA文库进行免疫筛选,鉴定出一个编码β-半乳糖苷结合凝集素(半乳糖凝集素)的cDNA。该cDNA序列具有串联重复结构,被命名为Hco-gal-2,与几种线虫的半乳糖凝集素以及哺乳动物4型半乳糖凝集素具有显著的相似性。使用乳糖-琼脂糖亲和层析,天然半乳糖凝集素优先从H-gal-GP复合物中提取,也从成虫制备的不溶性膜部分中提取。亲和纯化的物质在SDS-PAGE上的表观分子量约为35 kDa,可见3条不同的条带。在蛋白质免疫印迹中,通过与针对重组Hco-GAL-2产生的抗血清反应,所有3条带均被鉴定为半乳糖凝集素。为了确定捻转血矛线虫半乳糖凝集素是否具有任何抗感染保护能力,用亲和纯化的半乳糖凝集素对羔羊进行疫苗接种,随后进行单次攻击感染。通过粪便虫卵计数或虫体计数判断,疫苗接种并未赋予对捻转血矛线虫感染的任何保护作用。

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