PF3D7_1330600

PF3D7_1330600 (tufA) encodes the apicoplast-encoded elongation factor Tu (EF-Tu) of Plasmodium falciparum, a translationally active component of the organellar protein synthesis machinery whose product is localized within the apicoplast and maximally expressed in the trophozoite stage (PMID:15773980). As a canonical translation GTPase, recombinant PfEF-Tu hydrolyses GTP and undergoes nucleotide exchange (GDP release) mediated by the nuclear-encoded PfEF-Ts, with which it forms a functional complex (PMID:21163263). Beyond its translational role, PfEF-Tu displays an additional chaperone-related disulphide reductase activity, offering a rationale for retention of tufA on the plastid genome (PMID:21163263). The protein is a validated antimalarial target: it is directly bound and inhibited by structurally distinct EF-Tu antibiotics kirromycin, GE2270A, and enacyloxin IIa in vitro (PMID:10505418, PMID:21163263), and its levels are depleted by thiostrepton, which targets apicoplast translation (PMID:15773980).

1. 1999 Medium

   The first question was whether the apicoplast EF-Tu is a genuine drug target — reconstitution with recombinant protein showed it directly engages multiple classes of EF-Tu antibiotics that also kill parasites, establishing apicoplast translation as an antimalarial vulnerability.

   Evidence In vitro complex-formation assays with recombinant EF-Tu(pl) and kirromycin, GE2270A, enacyloxin IIa, plus blood-culture antimalarial assays

   PMID:10505418

   Open questions at the time

   • In vitro binding does not demonstrate that drug killing in culture proceeds through EF-Tu specifically
   • No enzymatic readout of inhibition at this stage
   • No structural mapping of the distinct binding sites
2. 2005 Medium

   It was unknown whether the plastid tufA gene is actually expressed as protein and where — localization and stage-specific analysis showed EF-Tu is produced and resides in the apicoplast with peak trophozoite expression, confirming an active organellar translation apparatus.

   Evidence Protein localization by immunofluorescence/fractionation and stage-specific expression analysis in P. falciparum

   PMID:15773980

   Open questions at the time

   • Does not define the full set of translation partners in the organelle
   • Stage-specific regulation mechanism not addressed
3. 2005 Medium

   To link a known antibiotic to apicoplast translation, thiostrepton treatment was shown to reduce EF-Tu protein levels, providing cellular evidence that this drug acts at the site of apicoplast protein synthesis.

   Evidence Drug treatment followed by western-blot quantification of EF-Tu protein levels in P. falciparum

   PMID:15773980

   Open questions at the time

   • Protein-level reduction does not distinguish direct from downstream effects on EF-Tu
   • Thiostrepton targets LSU rRNA, not EF-Tu itself
4. 2010 High

   The catalytic and partner biochemistry was resolved — reconstituted PfEF-Tu hydrolyses GTP, physically interacts with nuclear-encoded PfEF-Ts, and undergoes PfEF-Ts-mediated GDP release, establishing a functional EF-Tu/EF-Ts nucleotide-exchange cycle across the two genomes.

   Evidence Recombinant GTPase, nucleotide-exchange, and Co-IP/pulldown assays plus homology modeling of the PfEF-Tu:PfEF-Ts complex

   PMID:21163263

   Open questions at the time

   • Interaction with the ternary complex (aminoacyl-tRNA) not directly demonstrated
   • Complex geometry rests on homology modeling, not an experimental structure
5. 2010 Medium

   The antibiotic-target link was closed at the enzymatic level — kirromycin was shown to inhibit PfEF-Tu GTPase/translation activity in vitro, confirming EF-Tu as the direct molecular target rather than a bystander.

   Evidence In vitro enzymatic inhibition assay with recombinant PfEF-Tu and kirromycin

   PMID:21163263

   Open questions at the time

   • Enzymatic inhibition not yet tied to a defined resistance mutation in the parasite
   • Only kirromycin tested enzymatically among the inhibitor panel
6. 2010 Medium

   A moonlighting activity was uncovered — recombinant PfEF-Tu exhibits chaperone-related disulphide reductase activity, offering a functional explanation for why tufA is retained on the plastid genome.

   Evidence In vitro disulphide reductase biochemical assay with recombinant PfEF-Tu

   PMID:21163263

   Open questions at the time

   • Physiological substrates of the reductase activity not identified
   • In vivo relevance of this activity not tested

• Whether PfEF-Tu is genetically essential, how its stage-regulated expression is controlled, and the in vivo role and substrates of its disulphide reductase activity remain unresolved.
• No genetic knockout/knockdown of tufA in the corpus
• No experimental structure of PfEF-Tu or its complexes
• Resistance-conferring mutations to the antibiotic inhibitors not mapped