Plasmodium falciparum PhIL1-associated complex plays an essential role in merozoite reorientation and invasion of host erythrocytes
Fig 2
PhIL1-associated novel complex overlaps with components of the linear motor in the IMC but is a separate complex.
(A) Native PAGE analysis of schizont stage parasite lysate shows co-existence of PhIL1-associated complex with the glideosomal complex via. some overlapping components. A complex of ∼800 kDa was detected containing components of PfPhIL1-associated complex and GAP50. Furthermore, a smaller complex of ∼250 kDa was also detected that included PfPhIL1 and PfGAPM2. These results indicate a possible association of PhIL1-associated complex and glideosomal complex. n = 2 experiments. (B) PhIL1-associated complex co-exists with the glideosomal complex as seen by Glycerol gradient co-sedimentation analysis. Glycerol gradient fractionation of P. falciparum schizont extract using 5 to 45% Glycerol gradient followed by immunoblotting of the fractions using specific antibodies showed the co-sedimentation of PfGAP50, PfPhIL1, PfALV5, PfPhIP, and PfGAPM2 together in fractions 5 to 11, particularly, in fraction 9 corresponding to ∼250 kDa molecular mass. n = 2 experiments. (C) Recombinant GAP50 and BSA were subjected to SDS-PAGE followed by (D) far western analysis showing the interaction of recombinant PfGAP50 protein with PfPhIL1. GAP50 was denatured and renatured on the membrane followed by incubation with recombinant PhIL1 protein and probed with α-PhIL1 antisera which recognized a band of ~43kDa corresponding to the size of GAP50. BSA used as control protein did not show any interaction with PfPhIL1. (E) Model showing molecular motor and partially overlying proposed PhIL1-associated complex (dotted box) in the parasite IMC. Photosensitized 5-[125I] Iodonaphthalene-1-azide Labelled Protein-1 (PhIL1); PhIL1 Interacting Protein (PhIP); Glideosome-Associated Protein 40, 45 and 50 (GAP40, 45 and 50); Glideosome-associated protein with multiple membrane spans (GAPM); myosin-A (MyoA); Myosin-A Tail domain Interacting Protein (MTIP); Connector Protein between actin and protein of the merozoite surface (CP); Protein of the Merozoite surface (MzP); Parasite Plasma Membrane (PPM).