Department of Animal, Plant and Soil Science and Centre for Agri Bioscience (Agri Bio), La Trobe University, Melbourne, VIC, Australia.
Department of Biochemistry and Molecular Biology, Monash University, Clayton, VIC, Australia.
Methods Mol Biol. 2022;2442:475-515. doi: 10.1007/978-1-0716-2055-7_26.
Galectin-11 (LGALS-11) and galectin-14 (LGALS-14) are ruminant specific galectins, first reported in sheep. Although their roles in parasite immunity are still being elucidated, it appears that they influence protection against parasites. In gastrointestinal infections with the nematode Haemonchus contortus, both galectin-11 and galectin-14 appear to be protective. However, in a chronic infection of liver fluke, Fasciola hepatica, these galectins may aid parasite survival. To unravel the structural, functional, and ligand profile of galectin-11 and galectin-14, recombinant production of these proteins is vital. Here we present the recombinant production of soluble galectin-11 and galectin-14 from domestic sheep for in vitro and structural biology studies. These methods include parasite cultivation and infection, galectin staining of host and parasite tissue, surface staining of parasites with recombinant galectins, pull-down assays to identify endogenous galectin binding proteins, and in vitro assays to monitor the effect of galectins on parasite development.
半乳糖凝集素-11（LGALS-11）和半乳糖凝集素-14（LGALS-14）是反刍动物特有的半乳糖凝集素，最初在绵羊中报道。尽管它们在寄生虫免疫中的作用仍在阐明中，但它们似乎影响了对寄生虫的保护。在与线虫捻转血矛线虫的胃肠道感染中，半乳糖凝集素-11 和半乳糖凝集素-14 似乎都具有保护作用。然而，在慢性肝片吸虫感染中，这些凝集素可能有助于寄生虫的存活。为了解开半乳糖凝集素-11 和半乳糖凝集素-14 的结构、功能和配体特征，这些蛋白的重组生产至关重要。在这里，我们展示了从绵羊中重组生产可溶性半乳糖凝集素-11 和半乳糖凝集素-14，用于体外和结构生物学研究。这些方法包括寄生虫的培养和感染、宿主和寄生虫组织的半乳糖凝集素染色、重组半乳糖凝集素对寄生虫表面的染色、拉取实验以鉴定内源性半乳糖结合蛋白，以及体外实验以监测半乳糖凝集素对寄生虫发育的影响。
