Biomedical Engineering Reference
In-Depth Information
In addition to the success reported for the expression of semi-proplasmepsin
I using the thioredoxin fusion approach, the Rickey Yada's laboratory applied
the same approach to produce recombinant semi-proHAP and provided pre-
liminary characterization of the properties. 27 Two years later, Bhaumik et al. 46
utilized the preparation of this enzyme to obtain crystals of the mature enzyme
both in the apo form, where no inhibitor is bound, and in the complexes with
pepstatin and KNI-10006. The apo enzyme exhibited an unusual feature in that
zinc ions were observed to bind at several points on the enzyme surface, but
significantly one was observed binding to the active site His and Asp residues. It
is not thought that this metal binding is important in the catalytic mechanism
because Zn is absent in the complexes with the inhibitors. The pepstatin-HAP
structure is generally similar to those of other aspartic peptidases in complex
with pepstatin, but it was observed that an extremely large rigid body rotation
between the N- and C-terminal domains occurs compared to that seen for the
other enzymes when complexed with pepstatin.
The structure of the KNI-1006 complex requires special comments because,
as was seen in the plasmepsin I structure with this inhibitor, interactions are
seen that are not similar to the binding of peptstatin. In particular, the hydroxyl
of the inhibitor in the pepstatin complex binds between residues His and Asp as
expected; however, for KNI-10006, the hydroxyl group points away from those
residues. The interactions of the various components of the KNI compound do
not follow the 'canonical' binding in subsites along the active site cleft as seen
with pepstatin.
11.3.4 Plasmepsin IV
Plasmepsin IV is the fourth enzyme found in the digestive vacuole of P. falci-
parum. Wyatt and Berry described the preparation and properties of this
enzyme. 47 In particular, they showed that PfPMIV is able to cleave hemoglobin
at the Phe33-Leu34 bond known to be essential to initiate the degradation
process. In addition, this enzyme cleaves proteins found in the erythrocyte
membrane and shows strong activity on synthetic substrates. PfPMIV was
shown in this report to be inhibited by some of the classic inhibitors of the
family as well as some specific inhibitors developed for the purpose of blocking
hemoglobin digestion by the parasite. In 2003, Dame et al. published a report
on the genes for plasmepsin found in the four Plasmodium species infecting
man (P. falciparum, P. vivax, P. malariae, and P. ovalae), plus P. berghei,
P. knowlesi,andP. yoelii). 48 P. falciparum was the only species with as many as
10 plasmepsin genes; the other species do not exhibit genes for plasmepsins I, II,
and HAP but have the gene for plasmepsin IV.
Li et al. 49 subsequently expressed the P. falciparum gene encoding plasmepsin
IV as well as the genes from the other species infecting man. In an earlier report,
Westing et al. 50 had reported on studies of PfPMII, PvPM, and PmPM by
comparing substrate specificity and inhibitor binding; at that time, they were
unaware that the comparisons were between PfPMII, and the plasmepsins IV
from P. vivax and P. malariae.TheLiet al. report provided a detailed analysis of
Search WWH ::

Custom Search