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首页> 外文期刊>Infection and immunity >Expression of Cpgp40/15 in Toxoplasma gondii: a Surrogate System for the Study of Cryptosporidium Glycoprotein Antigens
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Expression of Cpgp40/15 in Toxoplasma gondii: a Surrogate System for the Study of Cryptosporidium Glycoprotein Antigens

机译：Cpgp40 / 15在弓形虫中的表达：隐孢子虫糖蛋白抗原研究的替代系统。

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Cryptosporidium parvum is a waterborne enteric coccidian that causes diarrheal disease in a wide range of hosts. Development of successful therapies is hampered by the inability to culture the parasite and the lack of a transfection system for genetic manipulation. The glycoprotein products of the Cpgp40/15 gene, gp40 and gp15, are involved in C. parvum sporozoite attachment to and invasion of host cells and, as such, may be good targets for anticryptosporidial therapies. However, the function of these antigens appears to be dependent on the presence of multiple O-linked α-N-acetylgalactosamine (α-GalNAc) determinants. A eukaryotic expression system that would produce proteins bearing glycosylation patterns similar to those found on the native C. parvum glycoproteins would greatly facilitate the molecular and functional characterization of these antigens. As a unique approach to this problem, the Cpgp40/15 gene was transiently expressed in Toxoplasma gondii, and the expressed recombinant glycoproteins were characterized. Antisera to gp40 and gp15 reacted with the surface membranes of tachyzoites expressing the Cpgp40/15 construct, and this reactivity colocalized with that of antiserum to the T. gondii surface protein SAG1. Surface membrane localization was dependent on the presence of the glycophosphatidylinositol anchor attachment site present in the gp15 coding sequence. The presence of terminal O-linked α-GalNAc determinants on the T. gondii recombinant gp40 was confirmed by reactivity with Helix pomatia lectin and the monoclonal antibody 4E9, which recognizes α-GalNAc residues, and digestion with α-N-acetylgalactosaminidase. In addition to appropriate localization and glycosylation, T. gondii apparently processes the gp40/15 precursor into the gp40 and gp15 component glycopolypeptides, albeit inefficiently. These results suggest that a surrogate system using T. gondii for the study of Cryptosporidium biology may be useful.
机译： Cryptosporidium parvum 是一种水性肠球菌，可在多种宿主中引起腹泻病。无法培养寄生虫以及缺乏用于基因操作的转染系统阻碍了成功疗法的发展。 Cpgp40 / 15 基因的糖蛋白产物gp40和gp15参与了 C。小孢子虫子体对宿主细胞的附着和侵袭，因此可能是抗隐孢子虫治疗的良好靶标。然而，这些抗原的功能似乎取决于多个O-连接的α- N -乙酰半乳糖胺（α-GalNAc）决定簇的存在。真核表达系统将产生带有糖基化模式的蛋白质，该蛋白质类似于天然C上发现的蛋白质。小分子糖蛋白将大大促进这些抗原的分子和功能表征。作为解决此问题的独特方法， Cpgp40 / 15 基因在弓形虫中瞬时表达，并表征了表达的重组糖蛋白。 gp40和gp15的抗血清与表达 Cpgp40 / 15 构建体的速殖子的表面膜反应，该反应性与抗血清对 T的反应共定位。弓形虫表面蛋白SAG1。表面膜的定位取决于在gp15编码序列中存在的糖磷脂酰肌醇锚附着位点的存在。在 T上存在末端O-连接的α-GalNAc决定簇。通过与 Helix pomatia 凝集素和识别α-GalNAc残基的单克隆抗体4E9的反应性，并用α- N -乙酰半乳糖苷酶消化，确认了冈地重组gp40。 。除了适当的定位和糖基化， T。刚地显然将gp40 / 15前体加工成gp40和gp15组分糖多肽，尽管效率很低。这些结果表明使用 T的替代系统。 gondii 用于研究隐孢子虫生物学可能是有用的。

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