Phosphoethanolamine-N-methyltransferase is a potential biomarker for the diagnosis of P. knowlesi and P. falciparum malaria

Abstract

BackgroundPlasmodium knowlesi is recognised as the main cause of human malaria in Southeast Asia. The disease is often misdiagnosed as P. falciparum or P. malariae infections by microscopy, and the disease is difficult to eliminate due to its presence in both humans and monkeys. P. knowlesi infections can rapidly cause severe disease and require prompt diagnosis and treatment. No protein biomarker exists for the rapid diagnostic test (RDT) detection of P. knowlesi infections. Plasmodium knowlesi infections can be diagnosed by PCR.Methods and principal findingsPhosphoethanolamine-N-methyltransferase (PMT) is involved in malaria lipid biosynthesis and is not found in the human host. The P. falciparum, P. vivax and P. knowlesi PMT proteins were recombinantly expressed in BL21(DE3) Escherichia coli host cells, affinity purified and used to raise antibodies in chickens. Antibodies against each recombinant PMT protein all detected all three recombinant proteins and the native 29 kDa P. falciparum PMT protein on western blots and in ELISA. Antibodies against a PMT epitope (PLENNQYTDEGVKC) common to all three PMT orthologues detected all three proteins. Antibodies against unique peptides from each orthologue of PMT, PfCEVEHKYLHENKE, PvVYSIKEYNSLKDC, PkLYPTDEYNSLKDC detected only the parent protein in western blots and P. falciparum infected red blood cell lysates or blood lysates spiked with the respective proteins. Similar concentrations of PfPMT and the control, PfLDH, were detected in the same parasite lysate. The recombinant PfPMT protein was detected by a human anti-malaria antibody pool.ConclusionPMT, like the pan-specific LDH biomarker used in RDT tests, is both soluble, present at comparable concentrations in the parasite and constitutes a promising antimalarial drug target. PMT is absent from the human proteome. PMT has the potential as a biomarker for human malaria and in particular as the first P. knowlesi specific protein with diagnostic potential for the identification of a P. knowlesi infection.

A Dual, Systematic Approach to Malaria Diagnostic Biomarker Discovery

“…Eleven candidates targeted P. falciparum , 10 P. vivax , 2 P. knowlesi , and 2 pan. Phosphoethanolamine N-methyltransferase (PMT) was evaluated for its ability to indicate P. falciparum , P. vivax , and P. knowlesi [ 37 ]. Aldolase (FBPA) made it onto the list of parasite proteins since the article we included in our review identified a P. vivax -specific version of this biomarker [ 47 ].…”

“…In addition, PMT met all but 1: expression by all human-infecting Plasmodium species. A P. malariae orthologue appears to be lacking in PlasmoDB, even though a putative orthologue is predicted to exist [ 37 ]. The studies describing these 3 candidates were classified as early (nonhuman) discovery studies with a level 2 deployability based on the enzyme-linked immunosorbent assay (ELISA) used for detection [ 28 , 37 , 51 ].…”

“…A P. malariae orthologue appears to be lacking in PlasmoDB, even though a putative orthologue is predicted to exist [ 37 ]. The studies describing these 3 candidates were classified as early (nonhuman) discovery studies with a level 2 deployability based on the enzyme-linked immunosorbent assay (ELISA) used for detection [ 28 , 37 , 51 ]. Conversely, merozoite surface protein 1 and rhoptry protein 3 possessed all of the favorable diagnostic features but did not reach the ring-stage abundance threshold established.…”

Exploring the use of Circumsporozoite Protein (CSP) as Diagnostic Biomarker for Specific Detection of Plasmodium Knowlesi

“…Thus, the speci city of the antibodies targeting this epitope would require further analysis. In Krause and Goldring (2018), it was demonstrated that phosphoethanolamine-Nmethyltransferase (PMT) may be able to be used as a target biomarker in malaria RDT [16]. The researchers identi ed PMT epitopes that are speci c to P. knowlesi, P. vivax, and P. falciparum in in silico analysis.…”

Computational and experimental elucidation of Plasmodium falciparum phosphoethanolamine methyltransferase inhibitors: Pivotal drug target

“…Therefore; the inhibition of catalytic domain of Pf PMT can lead to complete inhibition of SDPM pathway. Since Pf PMT has been found to be expressed throughout the asexual and sexual phases of parasite life cycle and absence of its human orthologues therefore it may be used as target template for development of new antimalarial [14–16]. Biochemical studies have demonstrated that human malaria parasite orthologs of PMT have similar structural as well as biochemical properties and inhibition profiles as plasmodium [17].…”