Inhibitory monoclonal antibodies recognise epitopes adjacent to a proteolytic cleavage site on the RAP-1 protein of Plasmodium falciparum

{ "0" : "Institute of Cell and Molecular Biology, Edinburgh, UK" , "1" : "Institute of Cell, Animal and Population Biology, University of Edinburgh, Edinburgh, UK" , "2" : "Max-Planck-Institut für Biochemie, Martinsried bei München, Germany" , "4" : "Malaria" , "5" : "Rhoptry" , "6" : "Inhibitory monoclonal antibody" , "7" : "RAP-1" , "8" : "Epitope mapping" , "9" : "mAb, monoclonal antibody" , "10" : "IPTG, isopropyl-β-thiogalactopyranoside" , "11" : "SDS, sodium dodecyl sulphate" , "12" : "PAGE, polyacrylamide gel electrophoresis" , "13" : "LB, Luria broth"}
Molecular and Biochemical Parasitology (Impact Factor: 2.73). 11/1992; DOI: 10.1016/0166-6851(92)90138-A

ABSTRACT The low-molecular-weight rhoptry-associated protein (RAP) complex of Plasmodium falciparum consists of at least two gene products, RAP-1 and RAP-2, and has the ability to immunise Saimiri monkeys against experimental P. falciparum infection. Several monoclonal antibodies specifically recognise this complex and in this study we show that purified immunoglobulin derived from these monoclonals is capable of inhibiting parasite growth in vitro. It has previously been shown that RAP-1 initially appears as an 80-kDa protein (p80) in early schizogony and is processed to a 65-kDa protein (p65) in late schizogony. Several of the inhibitory monoclonals recognise both the 80- and 65-kDa proteins by Western blot analysis suggesting that they recognise linear epitopes on RAP-1. We have mapped these epitopes by testing the reactivity of the monoclonals against fragments of the rap-1 gene expressed as β-galactosidase fusion proteins and subsequently against synthetic peptides. All of the epitopes map to a region 10–20 amino acids C-terminal to the proteolytic cleavage site for the processing of p80 to p65 at amino acid 190. We also show that the 65-kDa protein is not present in purified merozoites, suggesting that its generation is associated with merozoite release rather than erythrocyte invasion. These results are discussed with respect to possible inhibitory mechanisms for the monoclonals.

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