Computational and experimental studies of plasmodium falciparum protein PfAMA1 domain-II loop dynamics: implications in PfAMA1-PfRON2 binding event

Protein-protein interactions are interesting targets for various drug discovery campaigns. One such promising and therapeutically pertinent protein-protein complex is PfAMA1-PfRON2, which is involved in malarial parasite invasion into human red blood cells. A thorough understanding of the interactions between these macromolecular binding partners is absolutely necessary to design better therapeutics to fight against the age-old disease affecting mostly under-developed nations. Although crystal structures of several PfAMA1-PfRON2 complexes have been solved to understand the molecular interactions between these two proteins, the mechanistic aspects of the domain II loop-PfRON2 association is far from clear. The current work investigates a crucial part of the recognition event; i.e., how the domain II loop of PfAMA1 exerts its effect on the alpha helix of the PfRON2, thus influencing the overall kinetics of this intricate recognition phenomenon. To this end, we have conducted thorough computational investigation of the dynamics and free energetics of domain II loop closing processes using molecular dynamics simulation. The computational results are validated by systematic alanine substitutions of the PfRON2 peptide helix. The subsequent evaluation of the binding affinity of Ala-substituted PfRON2 peptide ligands by surface plasmon resonance (SPR) and isothermal titration calorimetry (ITC) provides a rank of the relative importance of the residues in context. Our combined (computational and experimental) investigation has revealed that the domain II loop of PfAMA1 is in fact responsible for arresting the PfRON2 molecule from egress, K2027 and D2028 of PfRON2 being the determinant residues for the capturing event. Our study provides a comprehensive understanding of the molecular recognition event between PfAMA1 and PfRON2, specifically in the post binding stage, which potentially can be utilized for drug discovery against malaria. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=95 SRC="FIGDIR/small/463826v1_ufig1.gif" ALT="Figure 1"> View larger version (37K): org.highwire.dtl.DTLVardef@a16597org.highwire.dtl.DTLVardef@14cc03org.highwire.dtl.DTLVardef@15e0e57org.highwire.dtl.DTLVardef@1ff8905_HPS_FORMAT_FIGEXP M_FIG C_FIG