Abstract
Apical membrane antigen-1 (AMA1) is a transmembrane protein present on the surface of merozoites that is thought to be involved in the process of parasite invasion of host erythrocytes. Although it is the target of a natural immune response that can inhibit invasion, little is known about the molecular mechanisms by which AMA1 facilitates the invasion process. In an attempt to identify peptides that specifically interact with and block the function of AMA1, a random peptide library displayed on the surface of filamentous phage was panned on recombinant AMA1 from Plasmodium falciparum. Three peptides with affinity for AMA1 were isolated, and characterization of their fine binding specificities indicated that they bind to a similar region on the surface of AMA1. One of these peptides was found to be a potent inhibitor of the invasion of P. falciparum merozoites into human erythrocytes. We propose that this peptide blocks interaction between AMA1 and a ligand on the erythrocyte surface that is involved in a critical step in malarial invasion. The identification and characterization of these peptide inhibitors now permit an evaluation of the essential requirements that are necessary for efficient neutralization of merozoite invasion by blocking AMA1 function.
Highlights
Apical membrane antigen-1 (AMA1) is a transmembrane protein present on the surface of merozoites that is thought to be involved in the process of parasite invasion of host erythrocytes
A dramatic enrichment of phage with affinity for the antigen was observed after the third round of panning (Fig. 1A). These pools of phage showed no binding to the irrelevant proteins BSA and the ring infected erythrocyte surface antigen (RESA) (Fig. 1B), but did bind to Plasmodium chabaudi AMA1 (PcAMA1), which shares 52% amino acid sequence identity with Plasmodium falciparum AMA1 (PfAMA1)
The binding activity was conferred by the displayed peptides because phage lacking a peptide (control (C)) and two phage clones picked at random from the unpanned peptide library were unable to bind to either PfAMA1 or PcAMA1 (Fig. 2A)
Summary
The precise steps involved in merozoite invasion are not well understood, Chitnis and Blackman (25) have put forward some suggestions for the possible roles of various merozoite surface antigens in the overall invasion process. These peptides were found to perform most of the functions of the native hormones such as receptor binding, dimerization, and downstream signaling leading to biological activity. One of the peptides that we have isolated binds to recombinant AMA1 and recognizes the native protein in malarial parasites Binding of this peptide to AMA1 was found to inhibit the merozoite invasion of host erythrocytes, and alanine scanning has defined a small set of amino acid side chains that are essential for AMA1 binding. These peptides represent defined reagents that will help explore the structure of AMA1 and illuminate its function within the parasite life cycle and may provide lead compounds for future therapies based on inhibition of AMA1 function
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