Molecular Characterization and Immuno-Reactivity Patterns of a Novel Plasmodium falciparum Armadillo-Type Repeat Protein, PfATRP.

Emmanuel Amlabu,Henrietta Mensah-Brown,Ojo-Ajogu Akuh,Evelyn Quansah,Prince B Nyarko,Reuben Ayivor-Djanie,Manfred Anim,Laty G Thiam,Philip Ilani,Grace Opoku,Julian C Rayner,Gordon A Awandare

doi:10.3389/fcimb.2020.00114

Abstract

Nearly half of the genes in the Plasmodium falciparum genome have not yet been functionally investigated. We used homology-based structural modeling to identify multiple copies of Armadillo repeats within one uncharacterized gene expressed during the intraerythrocytic stages, PF3D7_0410600, subsequently referred to as P. falciparum Armadillo-Type Repeat Protein (PfATRP). Soluble recombinant PfATRP was expressed in a bacterial expression system, purified to apparent homogeneity and the identity of the recombinant PfATRP was confirmed by mass spectrometry. Affinity-purified α-PfATRP rabbit antibodies specifically recognized the recombinant protein. Immunofluorescence assays revealed that α-PfATRP rabbit antibodies reacted with P. falciparum schizonts. Anti-PfATRP antibody exhibited peripheral staining patterns around the merozoites. Given the localization of PfATRP in merozoites, we tested for an egress phenotype during schizont arrest assays and demonstrated that native PfATRP is inaccessible on the surface of merozoites in intact schizonts. Dual immunofluorescence assays with markers for the inner membrane complex (IMC) and microtubules suggest partial colocalization in both asexual and sexual stage parasites. Using the soluble recombinant PfATRP in a screen of plasma samples revealed that malaria-infected children have naturally acquired PfATRP-specific antibodies.

Highlights

Malaria is a major global health problem that poses a threat to half of the world’s population
The P. falciparum Armadillo-Type Repeat Protein (PfATRP) gene (PF3D7_0410600) is a 3-exon gene located on chromosome 4 and encodes a 326-amino acid protein with a predicted molecular weight of 32 kDa
PfATRP is evolutionarily conserved across rodent and primate Plasmodium species (Figure 1B) and all orthologs have a positionally-conserved cysteine residue at the C-terminal end of the protein

Summary

Introduction

Malaria is a major global health problem that poses a threat to half of the world’s population. Erythrocyte invasion by the malaria parasite is a complicated and highly coordinated process that involves attachment and penetration mediated by a sophisticated network of parasite proteins discharged from two apical secretory organelles, the rhoptries and micronemes (Cowman et al, 2017) Another key invasion-associated organelle is the inner membrane complex (IMC), which contributes to the maintenance of cell morphology and rigidity (Aikawa et al, 1981; Meszoely et al, 1987; Kono et al, 2012), and plays a role in motility and invasion, by acting as an anchor for the actin-myosin motor that provides the pre-requisite force necessary for invasion processes (Soldati et al, 2004; Baum et al, 2006, 2008; Jones et al, 2006; Yeoman et al, 2011). Considering the future prospects in taking advantage of protein-protein interactions for the development of better diagnostic tools for malaria infection, we sought to functionally characterize PF3D7_0410600 protein especially that it may play an important role in parasite-specific processes or could be a potential biomarker

Methods

Results

Conclusion