Plasmodium falciparum is the parasite responsible for the most severe form of
malaria. Its increasing resistance to existing
antimalarials represents a major threat to human health and urges the development of new therapeutic strategies to fight
malaria. The
proteasome is a
protease complex essential in all eukaryotes. Accordingly, inhibition of the Plasmodium
20S proteasome is highly toxic for the parasite at all of its infective and developmental stages.
Proteasome inhibitors have
antimalarial potential both as curative and transmission blocking agents, but in order to have therapeutic application, they must specifically target the Plasmodium
proteasome and not its human counterpart. X-ray crystallography has been widely used to determine structures of yeast and mammalian 20S proteasomes with
ligands. However, crystallisation of the Plasmodium
proteasome is challenging, as only small quantities of the complex can be directly purified from the parasite. Furthermore, most X-ray structures of
proteasome-inhibitor complexes require soaking of crystals with high concentrations of
ligand, thus preventing analysis of inhibitor subunit specificity. Instead we chose to determine the Plasmodium falciparum
20S proteasome structure, in the presence of a new rationally designed parasite-specific inhibitor, by high-resolution electron cryo-microscopy and single particle analysis. The resulting map, at a resolution of about 3.6 Å, allows a direct molecular analysis of
inhibitor/enzyme interactions. Here we present an overview of this structure, and how it provides valuable information that can be used to assist in the design of improved
proteasome inhibitors with the potential to be developed as next-generation
antimalarial drugs.