Erythrocytic stage-dependent regulation of oligomerization of Plasmodium ribosomal protein P2.

The eukaryotic 60 S-ribosomal stalk consists of P0, P1, and P2 proteins, which associate in a pentameric structure (P1(2)-P0-P2(2)). The Plasmodium falciparum protein P2 (PfP2) appears to play nonribosomal roles. It gets exported to the infected erythrocyte (IE) surface at 30 h post-merozoite invasion (PMI), concomitant with extensive oligomerization. Here we present certain biophysical properties of PfP2. Recombinant P2 (rPfP2) protein showed SDS-resistant oligomerization, which could be significantly abolished under reducing conditions. However, the protein continued to oligomerize even when both cysteine residues were mutated, and with up to 40 amino acids (aa) deleted from the C-terminal end. CD analysis of P2 showed largely α-helical and random coil domains. The SDS- and DTT-resistant oligomerization was studied further as it occurred in a development-specific manner in Plasmodium. In a synchronized erythrocytic culture of P. falciparum, the PfP2 protein was detected as part of the ribosomal complex (∼96 kDa) at 18 and 30 h PMI, and was SDS sensitive. However, at 30 h, large amounts of SDS-sensitive aggregates of >600 kDa were also seen. At 30 h PMI, each of the parasites, IE cytosol and IE ghost contained 60-80-kDa PfP2 complexes, which resolved to a single 65-kDa species on SDS-PAGE. Tetramethylrhodamine-labeled rPfP2 protein exhibited DTT- and SDS-resistant oligomerization when treated with P. falciparum parasite extracts only from 24 to 36 h PMI, and multiple proteins appeared to be required for this oligomerization. Understanding the regulation of oligomerization of PfP2 may help in the elucidation of the novel structure-function relationship in the export of PfP2 to the red cell surface.