Biomedical Engineering Reference
In-Depth Information
A series of studies have now shown that the plasmepsins contained within the
digestive vacuole of P. falciparum are not completely essential for parasite
growth.
58-60
Omara-Opyene et al.
59
knocked out each of the four digestive vacuole
enzymes and found that although the resulting species grew at slower rates, par-
ticularly for PfPMI and PfPMIV knockouts, the genes are not essential. However,
the slower growth rates indicate that they might be necessary for parasite devel-
opment. Some abnormal morphology was observed in about a third of the cells
observed. These authors concluded at that time that because of the apparent
redundancy in the hemoglobin digestion mechanism, it might be necessary to
develop drugs that can inhibit two or more of the plasmepsins. Liu et al.
60
and
Bonilla et al.
58
constructed double gene knockouts to further study the role of
plasmepsins in the digestive vacuole. Their studies suggested that the reason for
the redundancy in P. falciparum may be the slower growth observed when one or
more plasmepsin genes are knocked out. Finally, Moura et al.
61
reported the
knockout of PfPMI, PfPMII, HAP, and PfPMIV. Again, the resulting mutant
was viable, although with reduced replication rates, and hemoglobin processing
seemed almost normal. This establishes that other enzymes in the digestive
vacuole, specifically cysteine andmetallo-proteases, are able to compensate for the
loss of the plasmepsins. The mutant was still sensitive to inhibitors of the aspartic
protease class of enzyme, indicating that one of the other plasmepsins may be the
actual target of the inhibitors. The next section will turn to those enzymes.
11.4 Non-Digestive Vacuole Plasmepsins
Which enzyme or enzymes are essential for the degradation process? Dame and
colleagues conducted knockout studies on all four genes, PfPM1, PfPM2,
HAP, and PfPM4, of P. falciparum. It appears that there is redundancy in the
enzymes of the parasite food vacuole, as any single knockout does not dra-
matically impair parasite growth. Thus, a correlation between inhibition of
PfPM2 and killing of parasites in culture does not imply that PfPM2 is an
essential enzyme. An inhibitor could be blocking more than one of the four
enzymes or could be blocking one of the non-food vacuole enzymes. As men-
tioned earlier, the completion of the P. falciparum genome sequence has
revealed that a total of 10 genes encoding aspartic peptidase-like enzymes are
present. In the following sections, information about the known properties of
non-digestive vacuole enzymes will be provided.
Some potential functions of the non-digestive vacuole enzymes were dis-
cussed in 2001 by Salmon et al.
62
In order for the new merozoites to exit an
infected erythrocyte, they must carry out proteolysis of cellular components. By
inhibiting cysteine proteases of the parasite, they were able to block exit at a
stage just before cell rupture. While their experiments reveal an important
function of cysteine protease in the exit process, complete details of cell rupture
remain to be discovered, and it is possible that additional proteases are involved
in further steps. Release of daughter merozoites occurs in a two-step fashion, as
shown in a study by Wickham et al.
63
The first step is the degradation of the
