Tryptophan-rich domains of Plasmodium falciparum SURFIN_4.2 and Plasmodium vivax PvSTP2 interact with membrane skeleton of red blood cell

Plasmodium falciparum dramatically alters the morphology and properties of the infected red blood cells (iRBCs). A large group of exported proteins participate in these parasite-host interactions occurring at the iRBC membrane skeleton. SURFIN_4.2 is one of iRBC surface protein that belongs to surface-associated interspersed protein (SURFIN) family. Although the intracellular tryptophan-rich domain (WRD) was proposed to be important for the translocation of SURFINs from Maurer’s clefts to iRBC surface, the molecular basis of this observation has yet to be defined. The WRDs of P. falciparum SURFIN proteins and their orthologous Plasmodium vivax subtelomeric transmembrane proteins (PvSTPs) show homology to the intracellular regions of PfEMP1 and Pf332, both of which are involved in RBC membrane skeleton interactions, and contribute to malaria pathology.

Two transfected lines expressing recombinant SURFINs (NTC-GFP and NTC-4.2WRD2-GFP) of the 3D7 sequence were generated by transfection in P. falciparum. In vitro binding assays were performed by using recombinant WRDs of SURFIN_4.2/PvSTP2 and inside-out vesicles (IOVs). The interactions between the recombinant WRDs of SURFIN_4.2/PvSTP2 with actin and spectrin were evaluated by the actin spin down assay and an enzyme-linked immunosorbent assay based binding assays, respectively.

The recombinant SURFINs (NTC-4.2WRD2-GFP), in which the second WRD from SURFIN_4.2 was added back to NTC-GFP, show diffused pattern of fluorescence in the iRBC cytosol. Furthermore, WRDs of SURFIN_4.2/PvSTP2 were found to directly interact with the IOVs of RBC, with binding affinities ranging from 0.26 to 0.68 μM, values that are comparable to other reported parasite proteins that bind to the RBC membrane skeleton. Further experiments revealed that the second WRD of SURFIN_4.2 bound to F-actin (K _d = 5.16 μM) and spectrin (K _d = 0.51 μM).