Critical glycosylated residues in exon three of erythrocyte glycophorin A engage Plasmodium falciparum EBA-175 and define receptor specificity.
Critical glycosylated residues in exon three of erythrocyte glycophorin A engage Plasmodium falciparum EBA-175 and define receptor specificity.
复制标题
红细胞血型糖蛋白 A 外显子三中的关键糖基化残基与恶性疟原虫 EBA-175 结合并定义受体特异性。
DOI：
10.1128/mbio.01606-14
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发表时间：
2014
期刊：
影响因子：
6.4
通讯作者：
Tolia,NirajH
中科院分区：
文献类型：
--
作者：
Salinas,NicholeD;Paing,MayM;Tolia,NirajH
红细胞侵入是疟疾发病机制中的重要步骤。恶性疟原虫红细胞结合样蛋白（EBL）家族识别红细胞上的血型糖蛋白（Glycophorins，Gp），在疟原虫入侵过程中的粘附中起着关键作用。然而，每种寄生虫配体特异性受体识别的分子基础仍然难以捉摸，如恶性疟原虫EBA-175（PfEBA-175）/GpA配体/受体对的情况。这在很大程度上是由于难以产生适当的糖基化和功能性受体。在这里，我们开发了一种表达系统，以产生重组糖基化和功能性GpA，以及突变和截短。我们确定了PfEBA-175参与的基本结合区域和决定因素，证明了这些决定因素是抑制寄生虫生长所必需的，并确定了介导PfEBA-175-GpA相互作用的重要聚糖。结果表明，PfEBA-175接合由外显子3编码的GpA的多个聚糖，并且聚糖的呈递可能是高亲合力结合所需的。由于剪接位点突变，GpB和GpE中外显子3的缺失赋予PfEBA-175对GpA的特异性识别。我们推测GpB和GpE可能是由于选择性压力而失去GpA中的PfEBA-175结合位点而产生的。这里描述的表达系统具有更广泛的应用，检查其他EBL成员重要的寄生虫入侵，以及其他病原体，识别血型糖蛋白。定义受体-配体相互作用中的关键结合决定簇的能力，以及遗传操纵糖基化受体的系统，为设计干扰寄生虫invasion. IMPORTANCEPlasmodiumfalciparum的干预措施开辟了新的途径。关键进入途径涉及恶性疟原虫EBA-175（PfEBA-175）识别RBC上的血型糖蛋白A（GpA）。尽管了解这种蛋白质-蛋白质相互作用，但特异性受体结合的完整机制尚不清楚。PfEBA-175识别GpA，但不能结合相关的RBC受体GpB或GpE。了解使PfEBA-175能够特异性识别GpA的必要元素对于开发特异性和有效的PfEBA-175抑制剂至关重要，这些抑制剂可以破坏宿主细胞入侵并有助于疟疾控制。在这里，我们描述了一种新的系统来生产和操纵宿主受体GpA。使用这个系统，我们探讨了GpA中的元素参与，从而为宿主细胞的入侵。这些研究对于了解配体和受体如何相互作用以及疟疾干预措施的未来发展具有重要意义。
Erythrocyte invasion is an essential step in the pathogenesis of malaria. The erythrocyte binding-like (EBL) family of Plasmodium falciparum proteins recognizes glycophorins (Gp) on erythrocytes and plays a critical role in attachment during invasion. However, the molecular basis for specific receptor recognition by each parasite ligand has remained elusive, as is the case with the ligand/receptor pair P. falciparum EBA-175 (PfEBA-175)/GpA. This is due largely to difficulties in producing properly glycosylated and functional receptors. Here, we developed an expression system to produce recombinant glycosylated and functional GpA, as well as mutations and truncations. We identified the essential binding region and determinants for PfEBA-175 engagement, demonstrated that these determinants are required for the inhibition of parasite growth, and identified the glycans important in mediating the PfEBA-175–GpA interaction. The results suggest that PfEBA-175 engages multiple glycans of GpA encoded by exon 3 and that the presentation of glycans is likely required for high-avidity binding. The absence of exon 3 in GpB and GpE due to a splice site mutation confers specific recognition of GpA by PfEBA-175. We speculate that GpB and GpE may have arisen due to selective pressure to lose the PfEBA-175 binding site in GpA. The expression system described here has wider application for examining other EBL members important in parasite invasion, as well as additional pathogens that recognize glycophorins. The ability to define critical binding determinants in receptor-ligand interactions, as well as a system to genetically manipulate glycosylated receptors, opens new avenues for the design of interventions that disrupt parasite invasion.IMPORTANCEPlasmodium falciparum uses distinct ligands that bind host cell receptors for invasion of red blood cells (RBCs) during malaria infection. A key entry pathway involves P. falciparum EBA-175 (PfEBA-175) recognizing glycophorin A (GpA) on RBCs. Despite knowledge of this protein-protein interaction, the complete mechanism for specific receptor engagement is not known. PfEBA-175 recognizes GpA but is unable to engage the related RBC receptor GpB or GpE. Understanding the necessary elements that enable PfEBA-175 to specifically recognize GpA is critical in developing specific and potent inhibitors of PfEBA-175 that disrupt host cell invasion and aid in malaria control. Here, we describe a novel system to produce and manipulate the host receptor GpA. Using this system, we probed the elements in GpA necessary for engagement and thus for host cell invasion. These studies have important implications for understanding how ligands and receptors interact and for the future development of malaria interventions.