EK

EK (PF3D7_1124600) is a cytoplasmic, ethanolamine-specific kinase of P. falciparum that catalyses the committed first step of the CDP-ethanolamine (Kennedy) branch of phospholipid biosynthesis, generating phosphoethanolamine from ethanolamine (PMID:3013685, PMID:19845508). It is biochemically and genetically distinct from the parasite's choline kinase, being separable by higher specific activity, heat stability, and selective inhibition by D-2-amino-1-butanol rather than hemicholinium-3 (PMID:3013685). Recombinant PfEK follows Michaelis-Menten kinetics with a Km for ethanolamine of ~476 µM, is competitively inhibited by the bis-thiazolium compound T3, localizes to the parasite cytoplasm, and increases in expression during erythrocytic development (PMID:19845508). In P. berghei the locus is refractory to knockout under conditions where control tagging succeeds, indicating EK activity is essential for asexual blood-stage growth (PMID:20478340). EK phosphorylation is promoted by PfCDPK7 downstream of a PfPI4K→PfCDPK7 signaling axis that governs phospholipid biosynthesis, becoming hypophosphorylated when PfCDPK7 is depleted (PMID:34866326).

1. 1986 High

   It was unknown whether the ethanolamine kinase activity in infected erythrocytes was a distinct enzyme or a side-activity of choline kinase; kinetic and inhibitor profiling established EK as a separate, ethanolamine-specific enzyme feeding phosphoethanolamine into phospholipid synthesis.

   Evidence Enzyme activity assays, kinetics, selective inhibitor profiling, and heat-inactivation in P. falciparum-infected erythrocyte homogenates

   PMID:3013685

   Open questions at the time

   • Enzyme identity in homogenates not yet tied to a specific gene
   • No subcellular localization
2. 2009 High

   The distinct EK activity was assigned to a defined recombinant protein, confirming its ethanolamine specificity, kinetic parameters, inhibitor sensitivity, cytoplasmic localization, and developmental expression pattern.

   Evidence Recombinant His6-PfEK expressed in E. coli, in vitro kinetics and competitive inhibition assays, antibody-based localization and stage analysis in P. falciparum

   PMID:19845508

   Open questions at the time

   • No structural model
   • Regulation of activity not yet addressed
3. 2010 Medium

   Whether EK activity is dispensable was tested by genetic disruption; the locus was refractory to knockout while control tagging succeeded, establishing EK as essential for asexual blood-stage development.

   Evidence Knockout attempts and GFP-tagging of Kennedy-pathway loci in P. berghei with fluorescence microscopy

   PMID:20478340

   Open questions at the time

   • Essentiality inferred from failure-to-knock-out rather than conditional depletion
   • Shown in P. berghei ortholog, not directly in P. falciparum
4. 2021 Medium

   How EK is regulated in vivo was unresolved; phosphoproteomics placed EK downstream of a PfPI4K→PfCDPK7 signaling axis, with PfCDPK7 promoting its phosphorylation as part of coordinated control of phospholipid biosynthesis.

   Evidence Quantitative phosphoproteomics in PfCDPK7-knockdown P. falciparum with phospholipid analysis

   PMID:34866326

   Open questions at the time

   • EK phosphosite not mutated to test functional consequence
   • Direct vs indirect phosphorylation by PfCDPK7 not distinguished
   • Effect of phosphorylation on EK catalytic activity unmeasured

• It remains unknown how EK phosphorylation controls its catalytic activity and whether EK is directly required in P. falciparum blood stages under conditional depletion, and no structural basis for its ethanolamine specificity or inhibitor binding has been defined.
• No structure of EK
• No conditional-knockdown phenotype in P. falciparum
• Functional link between phosphorylation and activity untested