trans

CARBOHYDRATE METABOLISM 159
assessed for the Plasmodium enzyme, but this
appears likely, given that Plasmodium PFK
exhibits greatest sequence similarity to that of
other apicomplexans. A primary dependence
on pyrophosphate could explain the observed
lack of nucleotide regulation, and would also
increase the efficiency of ATP production for
the entire glycolytic pathway by 50%.
A three-dimensional structure is available for
P. falciparum fructose bisphosphate aldolase,
providing the potential for structure-based
design of selective inhibitors. Although both
proteins are well conserved throughout evolution,
some structural differences are evident
in the Plasmodium enzymes. The C-terminal
tail of aldolase is critical for enzyme activity,
and displays significant divergence from the
human enzyme. Expression of antisense
oligonucleotides specific for parasite aldolase
has been reported to reduce enzyme activity,
slowing ATP production and inhibiting P. falciparum
growth in culture.
The crystal structure of triose phosphate
isomerase (TPI) has also been solved, revealing
significant differences from the human
enzyme, including the presence of different
amino acids adjacent to the catalytic residues
and at the homodimer interface. These structural
variations hold promise for development
of small molecule inhibitors at the active site
and/or peptide (or other inhibitors) that disrupt
the dimerization required for function of
this enzyme.
In keeping with the common themes noted
above, transcription of P. falciparum glyceraldehyde
3-phosphate dehydrogenase (GAPDH) is
induced 35-fold within the parasitized red
cell. Recombinant P. falciparum GAPDH specifically
reduces NAD rather than NADP (as
in the human enzyme), providing the NADH
necessary to drive lactate dehydrogenase
activity at the end of the glycolytic pathway
(see below).
The specific activity of Plasmodium phosphoglycerate
kinase (PGK) is greatest (7-fold
higher than host-cell PGK activity) during the
later phases of intraerythrocytic growth. Activity
is enhanced by the addition of a mild detergent,
supporting the concept that malarial glycolytic
enzymes may be associated with membranous
or cytoskeletal components of the cell,
as noted above.
All of the subsequent enzymes in the glycolytic
pathway – which constitute the ATPgenerating
steps – can be recognized in the
P. falciparum genome database, although only
enolase has been characterized in any detail.
Interestingly, both the GAPDH and enolase proteins
(of Plasmodium and other apicomplexans
as well) contain amino acid insertions and
deletions (indels) that are otherwise restricted
to organisms that harbor endosymbiotic plastid
organelles (Chapter 12). These amino acid
insertions provide valuable information concerning
the evolutionary relationship of apicomplexans
with plants, dinoflagellates, algae,
and other plastid-bearing species, and also
define variations in enzyme structure that may
be exploited for pharmacological intervention.
The pyruvate kinase (PK) of other apicomplexan
species, including Toxoplasma gondii,
Eimeria tenella and Cryptosporidium parvum,
has been characterized in some detail. Unlike
mammalian and other PK enzymes, the apicomplexan
enzyme is not significantly influenced
by fructose 1,6-bisphosphate, but is
strongly activated by the upstream monophosphate
derivatives of glucose and fructose. This
unusual regulation likely reflects the lack of
regulation at the PFK step, as noted above. As
a result, fructose 1,6-bisphosphate levels are
a poor indicator of glycolytic load in apicomplexan
parasites.
Rather than contributing to mitochondrial
processes, most of the NADH produced
by GAPDH is consumed by a potent lactate
BIOCHEMISTRY AND CELL BIOLOGY: PROTOZOA