RESUMO
The discovery of effective new antimalarial agents is urgently needed. One of the most frequently studied molecules anchored to the parasite surface is the merozoite surface protein-1 (MSP1). At red blood cell invasion MSP1 is proteolytically processed, and the 19-kDa C-terminal fragment (MSP119) remains on the surface and is taken into the red blood cell, where it is transferred to the food vacuole and persists until the end of the intracellular cycle. Because a number of specific antibodies inhibit erythrocyte invasion and parasite growth, MSP119 is therefore a promising target against malaria. Given the structural homology of cupredoxins with the Fab domain of monoclonal antibodies, an approach combining NMR and isothermal titration calorimetry (ITC) measurements with docking calculations based on BiGGER is employed on MSP119-cupredoxin complexes. Among the cupredoxins tested, rusticyanin forms a well defined complex with MSP119 at a site that overlaps with the surface recognized by the inhibitory antibodies. The addition of holo-rusticyanin to infected cells results in parasitemia inhibition, but negligible effects on parasite growth can be observed for apo-rusticyanin and other proteins of the cupredoxin family. These findings point to rusticyanin as an excellent therapeutic tool for malaria treatment and provide valuable information for drug design.
Assuntos
Antimaláricos/farmacologia , Azurina/metabolismo , Azurina/farmacologia , Proteína 1 de Superfície de Merozoito/metabolismo , Plasmodium yoelii/metabolismo , Motivos de Aminoácidos , Sequência de Aminoácidos , Anticorpos Monoclonais/química , Apoproteínas/metabolismo , Azurina/química , Calorimetria , Sequência Conservada , Fragmentos Fab das Imunoglobulinas/química , Espectroscopia de Ressonância Magnética , Proteína 1 de Superfície de Merozoito/química , Simulação de Acoplamento Molecular , Dados de Sequência Molecular , Oxirredução/efeitos dos fármacos , Plasmodium falciparum/efeitos dos fármacos , Plasmodium falciparum/crescimento & desenvolvimento , Plasmodium yoelii/efeitos dos fármacos , Ligação Proteica/efeitos dos fármacos , Alinhamento de Sequência , Software , TermodinâmicaRESUMO
Merozoite surface protein 1 (MSP1) has been identified as a target antigen for protective immune responses against asexual blood stage malaria, but effective vaccines based on MSP1 have not been developed so far. We have modified the sequence of Plasmodium yoelii MSP119 (the C-terminal region of the molecule) and examined the ability of the variant proteins to bind protective monoclonal antibodies and to induce protection by immunization. In parallel, we examined the structure of the protein and the consequences of the amino acid changes. Naturally occurring sequence polymorphisms reduced the binding of individual protective antibodies, indicating that they contribute to immune evasion, but immunization with these variant proteins still provided protective immunity. One variant that resulted in the localized distortion of a loop close to the N-terminus of MSP119 almost completely ablated protection by immunization, indicating the importance of this region of MSP119 as a target for protective immunity and in vaccine development.