FBPA

FBPA (PF3D7_1444800) is a class I fructose-1,6-bisphosphate aldolase that acts both as a glycolytic enzyme and as a structural bridge in the Plasmodium falciparum invasion motor (PMID:9521758, PMID:16321976). It is a single-copy gene product that forms a homotetramer, and its crystal structure diverges from human aldolase through a unique hydrophobic surface pocket and a reduced tetramer interface (PMID:9521758). A soluble, enzymatically active form localizes to the parasite cytoplasm, while an inactive membrane-associated form has also been detected (PMID:2190085). Beyond its catalytic role, FBPA physically links the actin-myosin motor to invasion adhesins: it binds the C-terminal cytoplasmic tails of the TRAP-family adhesins TRAP and MTRAP, of AMA1, and of members of the EBL and RH ligand families through an induced-fit mechanism in which an aromatic residue of the ligand tail inserts into a hydrophobic pocket, causing arginine rotamer changes and a helix-loop-helix subdomain shift (PMID:17426153, PMID:16321976, PMID:22991428, PMID:27607074). These adhesin tails compete for overlapping binding sites on FBPA, and phosphorylation of the MTRAP tail by CDPK1 and PKA increases FBPA binding affinity ~10-fold (PMID:27607074). FBPA simultaneously binds both G- and F-actin with high affinity and promotes actin polymerization, providing the second half of the mechanical link (PMID:25261592). The FBPA-TRAP interaction is functionally required for motility: a small molecule stabilizing the complex reduces liver-cell invasion by ~95% and blocks gliding (PMID:26289816).

1. 1990 Medium

   Establishing that FBPA is a genuine catalytic aldolase defined its baseline identity before any moonlighting role was known.

   Evidence gene cloning and expression of enzymatically active recombinant protein in E. coli, with cell fractionation showing cytoplasmic and membrane-associated forms

   PMID:2190085

   Open questions at the time

   • GPI-anchor-mediated membrane association inferred from phospholipase C digestion, not directly demonstrated
   • functional significance of the inactive membrane form unresolved
2. 1992 Medium

   Kinetic characterization showed the parasite aldolase is enzymatically distinct from mammalian isoenzymes, indicating a divergent enzyme rather than a host contaminant.

   Evidence recombinant expression and Km/Vmax kinetics of P. berghei aldolase isoenzymes with stage-specific immunodetection

   PMID:1625704

   Open questions at the time

   • performed on P. berghei isoenzymes, not directly on PF3D7_1444800
   • stage-specific expression pattern not mapped in P. falciparum
3. 1994 Medium

   Defining the pre-mRNA splicing that supplies the initiation codon clarified prior anomalous cDNA clones and the gene architecture of the aldolase locus.

   Evidence reverse-PCR cDNA sequencing, in vitro translation, and Northern blot in P. berghei aldo-2

   PMID:8008025

   Open questions at the time

   • shown for P. berghei aldo-2, not PF3D7_1444800
   • regulatory role of accumulated unspliced precursor unknown
4. 1998 High

   The crystal structure revealed parasite-specific features—a unique hydrophobic surface pocket and altered tetramer interface—establishing structural targets distinguishing FBPA from human aldolase.

   Evidence X-ray crystallography at 3.0 Å (homotetramer)

   PMID:9521758

   Open questions at the time

   • functional role of the unique surface pocket not addressed at this stage
   • no ligand bound
5. 2005 Medium

   Demonstrating FBPA binding to the MTRAP cytoplasmic tail positioned it as the physical bridge between an invasion adhesin and the actin-myosin motor in merozoites.

   Evidence in vitro binding assay of MTRAP cytoplasmic tail to aldolase plus co-localization

   PMID:16321976

   Open questions at the time

   • binding not yet linked to actin at structural resolution
   • in vivo requirement not tested
6. 2007 High

   Co-crystallization with the TRAP tail defined the induced-fit binding mechanism and revealed that the adhesin- and actin-binding sites overlap, explaining how one aldolase could couple two partners.

   Evidence X-ray structures at 2.4 and 2.7 Å of FBPA with TRAP hexapeptide plus site-directed mutagenesis

   PMID:17426153

   Open questions at the time

   • actin binding at the overlapping site inferred, not co-crystallized
   • consequence for catalytic activity during binding not defined
7. 2012 High

   Extending the binding repertoire to RH-family ligand tails defined the sequence rules (an aromatic residue flanked by acidic residues) and showed selectivity among ligands.

   Evidence surface plasmon resonance, 2.5 Å crystal structure with RH2b peptide, aromatic-residue analysis

   PMID:22991428

   Open questions at the time

   • basis for selectivity (RH2b vs RH2a, DBL exclusion) not fully mechanistically explained
   • in vivo relevance of each ligand interaction untested
8. 2014 High

   Quantifying high-affinity binding to both G- and F-actin and showing FBPA promotes actin polymerization established that FBPA actively contributes to the actin half of the motor linkage.

   Evidence biolayer interferometry, co-sedimentation, and actin polymerization assays

   PMID:25261592

   Open questions at the time

   • structural basis of actin binding not resolved
   • polymerization effect not tested in intact motor context
9. 2015 High

   A small molecule stabilizing the FBPA-TRAP complex blocking invasion provided direct functional proof that this protein-protein interaction is required for motility and invasion.

   Evidence virtual screening, ternary FBPA-TRAP-compound 24 crystal structure, gliding and liver-cell invasion assays (~95% reduction)

   PMID:26289816

   Open questions at the time

   • genetic ablation of the interaction not performed
   • distinction between glycolytic and scaffold contributions to the phenotype not resolved
10. 2016 High

    Mapping the full adhesin binding set, defining CTD sequence requirements, and demonstrating phosphorylation-enhanced binding integrated FBPA into a regulatable hub coupling multiple ligands to actin.

    Evidence quantitative binding, CTD mutagenesis, CDPK1/PKA kinase assays, competition assays, and in-parasite MTRAP phosphorylation analysis

    PMID:27607074

    Open questions at the time

    • MTRAP phosphorylation not detected in asexual blood-stage parasites, leaving the in vivo trigger unresolved
    • physiological hierarchy among competing ligands unknown

• Whether the glycolytic and scaffolding functions of FBPA can be genetically separated, and how binding is regulated during native invasion, remains open.
• no genetic separation-of-function mutant reported
• in vivo phosphorylation trigger for enhanced binding not established
• functional role of the membrane-associated inactive form unknown