A targetable receptor-binding site on PfCyRPA to aid in the fight against malaria

Abstract

Recently, Day et al. https://doi.org/10.1016/j.celrep.2024.114012 identify a receptor binding site on the malaria parasite protein PfCyRPA that binds the host sugar Neu5Ac, and they find that disrupting this interaction impedes parasite growth. A map of the receptor-binding site identifies an attractive target for anti-malarial vaccines and therapeutics.

Malaria caused by the parasite Plasmodium falciparum remains one of the deadliest diseases, killing more than 600,000 people per year. Symptomatic infection occurs during the blood-stage of infection, when the merozoite stage of the parasite invades and reproduces in red blood cells. The parasite proteins and host receptors that mediate invasion have been the subject of intense study for decades because they provide some of the most promising targets for vaccines and therapeutics [1]. However, the numerous, and often redundant, interactions employed by Plasmodium falciparum have impeded our understanding of the invasion process and the development of countermeasures.

The five-component PfPTRAMP-CSS-RIPR-CyRPA-Rh5 (PCRCR) complex has recently gained attention because it is essential for merozoite invasion, but the function of each component has not yet been elucidated (Figure 1) [2,3]. Rh5 is the apical member of the complex and directly binds the host receptor basigin. Antibodies that block the Rh5-basigin interaction prevent parasite invasion, and Rh5 is a leading malaria vaccine candidate. RIPR has also been reported to bind the host receptor SEMA7A [4]. The other members of the PCRCR complex have often been depicted as simple linkers that connect Rh5 to the parasite membrane. However, antibodies that target the other members of the PCRCR complex also prevent parasite invasion, hinting at more complex functions for these proteins.

Figure 1. CyRPA is a glycan-binding member of the PCRCR complex.

Day et al. discovered a glycan binding site (red) on the merozoite protein CyRPA (orange) [5]. Blocking this interaction can inhibit parasite invasion of erythrocytes and could be the target of vaccines and therapeutics. A composite model of CyRPA in the context of the PCRCR complex and the host receptor basigin was assembled using existing structures of basigin (pink), Rh5 (cyan), RIPR (yellow), CSS (green), and an alphafold prediction of PTRAMP (grey). The unknown host protein carrying the glycan is shown in white outline. This figure was created using BioRender.

In a recent study, Day et al. revealed a novel and essential host-parasite interaction between CyRPA and α2–6-linked N-acetylneuraminic acid (Neu5Ac) [5]. The structure of CyRPA is similar to sialidase proteins but it lacks catalytic residues. This observation motivated Day et al. to perform a glycan-binding screen followed by quantitative binding experiments that demonstrated a preference for glycans terminating in Neu5Ac. A biantennary glycan with α2–6-linked Neu5Ac binds with particularly high affinity. Interestingly, the Rh5 receptor, basigin, and the closely associated CD44 protein exhibit this glycosylation pattern and knockout of these proteins impedes CyRPA binding. This suggests the formation of a large and intricate host-parasite protein complex that will be a fascinating subject of future studies.

Day et al. also determined the glycan binding site on CyRPA and used mutagenesis to validate the functional importance of this interaction [5]. Molecular modeling identified two potential Neu5Ac binding sites and mutagenesis at these sites disrupted glycan binding in vitro. These results support the preference for biantennary ligands and map the location of binding. Furthermore, a point mutation in one of these pockets disrupts the growth of parasites, demonstrating that this interaction is functionally important, and confirming that this interaction is a potential therapeutic target.

The location of the glycan-binding site explains the mechanism of several anti-CyRPA neutralizing monoclonal antibodies (mAbs). Many anti-CyRPA mAbs are neutralizing and their epitopes have been mapped, however the mechanistic basis for this neutralization has been a mystery [6–8]. The glycan binding site overlaps with the epitopes of several neutralizing mAbs, and Day et al. showed that these mAbs can prevent glycan binding. This further supports the use of the glycan binding site as a target for countermeasures. Interestingly, there are also neutralizing mAbs that do not block Neu5Ac binding, suggesting that there may still be more to learn about CyRPA function.

The receptor binding site mapped by Day et al. provides a valuable roadmap for the future development of malaria vaccines. CyRPA is an established blood-stage vaccine candidate that is more easily produced than Rh5 but may elicit antibodies with slightly lower inhibitory activity [9]. Day et al. showed that mAbs targeting the Rh5-interacting surface on CyRPA are non-neutralizing and mAbs targeting the Neu5Ac receptor-binding site are neutralizing.

Structural vaccinology can now be used to design a CyRPA vaccine that focuses the antibody response to neutralizing epitopes, such as the glycan-binding site, resulting in a potent and readily manufactured malaria vaccine.