Molecular Identification of a Malaria Merozoite Surface Sheddase

Abstract

Proteolytic shedding of surface proteins during invasion by apicomplexan parasites is a widespread phenomenon, thought to represent a mechanism by which the parasites disengage adhesin-receptor complexes in order to gain entry into their host cell. Erythrocyte invasion by merozoites of the malaria parasite Plasmodium falciparum requires the shedding of ectodomain components of two essential surface proteins, called MSP1 and AMA1. Both are released by the same merozoite surface “sheddase,” but the molecular identity and mode of action of this protease is unknown. Here we identify it as PfSUB2, an integral membrane subtilisin-like protease (subtilase). We show that PfSUB2 is stored in apical secretory organelles called micronemes. Upon merozoite release it is secreted onto the parasite surface and translocates to its posterior pole in an actin-dependent manner, a trafficking pattern predicted of the sheddase. Subtilase propeptides are usually selective inhibitors of their cognate protease, and the PfSUB2 propeptide is no exception; we show that recombinant PfSUB2 propeptide binds specifically to mature parasite-derived PfSUB2 and is a potent, selective inhibitor of MSP1 and AMA1 shedding, directly establishing PfSUB2 as the sheddase. PfSUB2 is a new potential target for drugs designed to prevent erythrocyte invasion by the malaria parasite.

A novel Plasmodium-specific prodomain fold regulates the malaria drug target SUB1 subtilase

“…SUB1 executes the specific processing of proteins involved in the egress of Plasmodium merozoites (Mz) from host hepatocytes or erythrocytes 11,14 , as well as Mz protein surface maturation 15 . SUB2 carries out processing of proteins involved in Mz invasion into erythrocytes 8,16 . This key multi-step process is considered the Achille's heal of the parasite life cycle and a promising vaccine and/or drug target 17 .…”

Fine Mapping of an Epitope Recognized by an Invasion-inhibitory Monoclonal Antibody on the Malaria Vaccine Candidate Apical Membrane Antigen 1

“…Upon schizont rupture PfAMA1 66 (but not its PfAMA1 83 precursor) is released from the micronemes to distribute across the surface of the free merozoite (38,39). At or around the point of invasion the molecule is then quantitatively shed by a membrane-bound subtilisin-like "sheddase" called PfSUB2, which translocates across the parasite surface, possibly in association with the moving junction (40,41). Shedding is mediated by a single cleavage just 29 residues distal to the transmembrane domain, resulting in release of essentially the entire AMA1 ectodomain in the form of a 48-kDa polypeptide called PfAMA1 48 ; an additional low level processing event within this molecule generates two disulfide-linked fragments, the larger of which is called PfAMA1 44 (41).…”

Cryptosporidium parvum glycoprotein gp40 localizes to the sporozoite surface by association with gp15

“…MSP1 is a GPI anchored 190kDa glycoprotein that is cleaved into several polypeptides that non-covalently associate with each other [15,16]. The 42 kDa C-terminal domain of MSP1 is a substrate for PfSUB2 [17], a serine protease that may be similar to the furin-like serine protease that cleaves gp40/15 [11]. Cleavage of the 42kDa fragment is necessary for merozoite invasion into erythrocytes [18].…”