Biochemical, biophysical, and functional characterization of bacterially expressed and refolded receptor binding domain of Plasmodium vivax Duffy-binding protein

Invasion of erythrocytes by malaria parasites is mediated by specific molec ular interactions. Plasmodium vivax is completely dependent on interaction with the Duffy blood group antigen to invade human erythrocytes, The P, viv ax Duffy-binding protein, which binds the Duffy antigen during invasion, be longs to a family of erythrocyte-binding proteins that also includes Plasmo dium falciparum sialic acid binding protein and Plasmodium knowlesi Duffy b inding protein, The receptor binding domains of these proteins lie in a con served, N-terminal, cysteine-rich region, region II, found in each of these proteins. Here, we have expressed P. vivax region II (PvRII), the P. vivax Duffy binding domain, in Escherichia coli, Recombinant PvRII is incorrectl y folded and accumulates in inclusion bodies. We have developed methods to refold and purify recombinant PvRII in its functional conformation. Biochem ical, biophysical, and functional characterization confirms that recombinan t PvRII is pure, homogeneous, and functionally active in that it binds Huff y-positive human erythrocytes with specificity. Refolded PvRII is highly im munogenic and elicits high titer antibodies that can inhibit binding of P. vivax Huffy-binding protein to erythrocytes, providing support for its deve lopment as a vaccine candidate for P. vivax malaria. Development of methods to produce functionally active recombinant PvRII is an important step for structural studies as well as vaccine development.