Molecular Parasitology Laboratory, University of Galway, Galway, Republic of Ireland.
Centre for Chromosome Biology, School of Natural Science, University of Galway, H91 TK33 Galway, Ireland.
PLoS Negl Trop Dis. 2024 Aug 30;18(8):e0012069. doi: 10.1371/journal.pntd.0012069. eCollection 2024 Aug.
Enolase is a 47 kDa enzyme that functions within the glycolysis and gluconeogenesis pathways involved in the reversible conversion of D-2-phosphoglycerate (2PGA) to phosphoenolpyruvate (PEP). However, in the context of host-pathogen interactions, enolase from different species of parasites, fungi and bacteria have been shown to contribute to adhesion processes by binding to proteins of the host extracellular matrix (ECM), such as fibronectin (FN) or laminin (LM). In addition, enolase is a plasminogen (PLG)-binding protein and induces its activation to plasmin, the main protease of the host fibrinolytic system. These secondary 'moonlighting' functions of enolase are suggested to facilitate pathogen migration through host tissues. This study aims to uncover the moonlighting role of enolase from the parasite Fasciola hepatica, shedding light on its relevance to host-parasite interactions in fasciolosis, a global zoonotic disease of increasing concern. A purified recombinant form of F. hepatica enolase (rFhENO), functioning as an active homodimeric glycolytic enzyme of ~94 kDa, was successfully obtained, fulfilling its canonical role. Immunoblotting studies on adult worm extracts showed that the enzyme is present in the tegument and the excretory/secretory products of the parasite, which supports its key role at the host-parasite interface. Confocal immunolocalisation studies of the protein in newly excysted juveniles and adult worms also localised its expression within the parasite tegument. Finally, we showed by ELISA that rFhENO can act as a parasitic adhesin by binding host LM, but not FN. rFhENO also binds PLG and enhances its conversion to plasmin in the presence of the tissue-type and urokinase-type PLG activators (t-PA and u-PA). This moonlighting adhesion-like function of the glycolytic protein enolase could contribute to the mechanisms by which F. hepatica efficiently invades and migrates within its host and encourages further research efforts that are designed to impede this function by vaccination or drug design.
烯醇酶是一种 47kDa 的酶，在糖酵解和糖异生途径中发挥作用，该途径涉及 D-2-磷酸甘油酸（2PGA）到磷酸烯醇丙酮酸（PEP）的可逆转化。然而，在宿主-病原体相互作用的背景下，来自不同种寄生虫、真菌和细菌的烯醇酶已被证明通过与宿主细胞外基质（ECM）中的蛋白质结合，如纤连蛋白（FN）或层粘连蛋白（LM），有助于黏附过程。此外，烯醇酶是纤溶酶原（PLG）结合蛋白，可诱导其激活为纤溶酶，即宿主纤维蛋白溶解系统的主要蛋白酶。这些烯醇酶的次要“兼职”功能被认为有助于病原体穿过宿主组织迁移。本研究旨在揭示寄生虫 Fasciola hepatica 烯醇酶的兼职作用，阐明其在 Fasciolosis 中与宿主-寄生虫相互作用的相关性，Fasciolosis 是一种日益受到关注的全球性人畜共患疾病。成功获得了纯化的重组 Fasciola hepatica 烯醇酶（rFhENO）形式，其作为~94kDa 的活性同源二聚体糖酵解酶发挥作用，满足了其典型作用。成虫提取物的免疫印迹研究表明，该酶存在于寄生虫的皮层和排泄/分泌产物中，这支持了其在宿主-寄生虫界面的关键作用。对刚孵出的幼虫和成虫中该蛋白的共聚焦免疫定位研究也将其表达定位于寄生虫皮层内。最后，我们通过 ELISA 表明 rFhENO 可以作为寄生虫黏附素通过与宿主 LM 结合发挥作用，但不能与 FN 结合。rFhENO 还与 PLG 结合，并在组织型和尿激酶型 PLG 激活剂（t-PA 和 u-PA）存在下增强其向纤溶酶的转化。糖酵解蛋白烯醇酶的这种兼职黏附样功能可能有助于 Fasciola hepatica 有效入侵和在其宿主内迁移的机制，并鼓励进一步研究，旨在通过疫苗接种或药物设计来阻止这种功能。
