The 190-kDa merozoite surface protein 1 (MSP-1) of Plasmodium falciparum, an essential component in the parasite's life cycle, is a primary candidate for a malaria vaccine. Rabbit antibodies elicited by the heterologously produced MSP-1 processing products p83, p30, p38, and p42, derived from strain 3D7, were analyzed for the potential to inhibit in vitro erythrocyte invasion by the parasite and parasite growth. Our data show that (i) epitopes recognized by antibodies, which inhibit parasite replication, are distributed throughout the entire MSP-1 molecule; (ii) when combined, antibodies specific for different regions of MSP-1 inhibit in a strictly additive manner; (iii) anti-MSP-1 antibodies interfere with erythrocyte invasion as well as with the intraerythrocytic growth of the parasite; and (iv) antibodies raised against MSP-1 of strain 3D7 strongly cross-inhibit replication of the heterologous strain FCB-1. Accordingly, anti-MSP-1 antibodies appear to be capable of interfering with parasite multiplication at more than one level. Since the overall immunogenicity profile of MSP-1 in rabbits closely resembles that found in sera of Aotus monkeys immunized with parasite-derived MSP-1 and of humans semi-immune to malaria from whom highly inhibiting antigen-specific antibodies were recovered, we consider the findings reported here to be relevant for the development of MSP-1-based vaccines against malaria.

Merozoites of the malaria parasite Plasmodium falciparum expose at their surface a large multiprotein complex, composed of proteolytically processed, noncovalently associated products of at least three genes, msp-1, msp-6, and msp-7. During invasion of erythrocytes, this complex is shed from the surface except for a small glycosylphosphatidylinositol-anchored portion originating from MSP-1. The proteolytic cleavage separating the C-terminal portion of MSP-1 is required for successful invasion. Little is known about the structure and function of the abundant and essential multipartite complex. Using heterologously produced MSP-1, MSP-6, and MSP-7 in precursor and with the exception of MSP-7 in processed form, we have studied in vitro the complex formation between the different proteins to identify the interaction partners within the complex. Both MSP-6 36 and MSP-7 bind only to MSP-1 subunits that are shed, but although MSP-6 36 contacts just subunit p38, MSP-7 interacts with p83, p30, and p38. The intact C-terminal region of MSP-6 is required for the association with p38 as well as for its multimerization into tetramers. Furthermore, our data suggest that only the processed form and not the precursor form of MSP-1 interacts with MSP-6 36 . MSP-6-as well as MSP-7-specific rabbit antibodies inhibit parasite multiplication in vitro as shown previously for antibodies directed against MSP-1. Our findings raise interesting questions with regard to proteolysis-mediated mechanisms of maturation of the MSP-1-MSP-6-MSP-7 complex and to the mode by which antibodies directed against this complex interfere with parasite multiplication.

The major protein component at the surface of merozoites, the infectious form of blood stage malaria parasites, is the merozoite surface protein 1 (MSP-1) complex. In the human malaria parasite Plasmodium falciparum, this complex is generated by proteolytic cleavage of a 190-kDa glycosylphosphatidylinositol-anchored precursor into four major fragments, which remain non-covalently associated. Here, we describe the in vitro reconstitution of the MSP-1 complex of P. falciparum strain 3D7 from its heterologously produced subunits. We provide evidence for the arrangement of the subunits within the complex and show how they interact with each other. Our data indicate that the conformation assumed by the reassembled complex as well as by the heterologously produced 190-kDa precursor corresponds to the native one. Based on these results we propose a first structural model for the MSP-1 complex. Together with access to faithfully produced material, this information will advance further structure-function studies of MSP-1 that plays an essential role during invasion of erythrocytes by the parasite and that is considered a promising candidate for a malaria vaccine.Merozoites, the erythrocyte invading form of malaria parasites, uniformly expose at their surface a major protein complex, the merozoite surface protein 1 (MSP-1).1 In Plasmodium falciparum, the parasite causing the most severe form of malaria in humans, MSP-1 is synthesized as an approximately 190-kDa precursor protein, which is deposited at the surface of the developing merozoite via a glycosylphosphatidylinositol anchor (1). In late schizogony, merozoites undergo maturation during which MSP-1 is proteolytically cleaved into four major fragments (2), which, however, remain non-covalently associated at the surface of the parasite. At the time of erythrocyte invasion, a second proteolytic cleavage separates the approximately 10-kDa glycosylphosphatidylinositol-anchored C terminus of MSP-1, called p19, from the rest of the complex, and only the membrane-bound portion is transferred into the newly infected erythrocyte while the remaining complex is shed from the surface of the parasite (3, 4).Several lines of evidence demonstrate that MSP-1 plays an essential role in the life cycle of the parasite and that it is involved in the erythrocyte invasion process. Thus, attempts to knock out the msp-1 gene in P. falciparum via homologous recombination failed, whereas the same approach allows to replace the functionally conserved C-terminal end of the molecule by a sequence of a distantly related Plasmodium species (5). Moreover, several monoclonal antibodies directed to the C-terminal p19 fragment inhibit efficiently erythrocyte invasion in vitro (6). These antibodies also prevent the secondary cleavage of MSP-1 during invasion (7). At least, this latter proteolytic step appears, therefore, to be an essential prerequisite for the infection of erythrocytes. Interestingly, antibodies were identified that can block the effect of invasion inhibiting antibodies in vitro (8). H...

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