The pathogenicity of Plasmodium falciparum is partly due to parasite-induced host cell modifications. These modifications are facilitated by exported P. falciparum proteins, collectively referred to as the exportome. Export of several hundred proteins is mediated by the PEXEL/HT, a protease cleavage site. The PEXEL/HT is usually comprised of five amino acids, of which R at position 1, L at position 3 and E, D, or Q at position 5 are conserved and important for export. Non-canonical PEXEL/HTs with K or H at position 1 and/or I at position 3 are presently considered non-functional. Here, we show that non-canonical PEXEL/HT proteins are overrepresented in P. falciparum and other Plasmodium species. Further, we show that non-canonical PEXEL/HTs can be cleaved and can promote export in both a REX3 and a GBP reporter, but not in a KAHRP reporter, indicating that non-canonical PEXEL/HTs are functional in concert with a supportive sequence environment. We then selected P. falciparum proteins with a non-canonical PEXEL/HT and show that some of these proteins are exported and that their export depends on non-canonical PEXEL/HTs. We conclude that PEXEL/HT plasticity is higher than appreciated and that non-canonical PEXEL/HT proteins cannot categorically be excluded from Plasmodium exportome predictions.
