trans

THE EMBDEN–MEYERHOF–PARNAS (EMP) PATHWAY OF GLYCOLYSIS 149
every trypanosomatid tested – T. congolense,
T. vivax, T. cruzi, Phytomonas, Crithidia and
Leishmania – with a good correlation between
the amount of protein and enzymatic activity.
The role of PPDK in glycosomes is not yet
clear. In addition to PPDK, there are at least four
other enzymes in the glycosome that produce
PP i : fatty acyl-CoA synthetase, hypoxanthineguanine
phosphoribosyl transferase, xanthine
phosphoribosyl transferase and orotate phosphoribosyl
transferase (Chapter 9). Each of
them catalyzes reactions with a G 0 close to
zero and thus PP i needs to be hydrolysed in
order to assure a continuation of the phosphoribosyl
transferase reactions of the purine
salvage and pyrimidine biosynthesis pathways.
While a pyrophosphatase activity has been
found in the cytosol of T. brucei, such activity
could not be detected in its glycosomes, and
this suggests that trypanosomes may have
developed an alternative route to hydrolyse
glycosomal PP i . PPDK could do so by converting
PEP into pyruvate while producing ATP.
Whether PPDK does in fact contribute in this
way to the glycosomal PP i and ATP balance
remains to be determined. The fact that PPDK
is not present in bloodstream-form glycosomes
would also suggest that the other four PP i -
producing enzymes may be absent from
bloodstream-form glycosomes as well. So far
only the absence of fatty-acid oxidation activity
from bloodstream-form trypanosomes has
been confirmed experimentally.
The presence of both a glycosomal ATPdependent
PFK with PP i -dependent characteristics
(see above), together with a
PP i -dependent PPDK, suggests that an ancestral
trypanosomatid may have possessed a
PP i -dependent type of glycolysis, similar to
what is still encountered in some anaerobic
protists, such as Giardia and Entamoeba
(Chapter 7, part 1). A subsequent adaptation
to an aerobic type of metabolism, where PFK
changed its phospho-substrate specificity
from PP i to ATP and where PPDK was replaced
by PYK for the conversion of PEP into pyruvate,
may have resulted in the replacement of
a PP i -dependent glycolysis by a completely
ATP-dependent one. PPDK would then have
been retained after having acquired its new
function in maintaining the PP i and ATP balance
inside glycosomes.
Since PPDK does not exist in higher eukaryotes,
the enzyme could be considered as a good
target for the design of drugs against Leishmania
and T. cruzi, provided that this enzyme
were essential for cell viability in these trypanosomatids.
The enzyme from T. brucei,
which has been crystallized and the structure of
which is being solved, would be a less appropriate
target, because it is not expressed in the
bloodstream form.
The hexose-monophosphate pathway
Glucose 6-phosphate (G6P) is also metabolized
by the hexose-monophosphate pathway (HMP),
also known as the pentose-phosphate shunt
(Figure 7.5). While the role of glycolysis is the
generation of ATP and pyruvate, the HMP is
mainly involved in maintaining a pool of cellular
NADPH, which may serve as a hydrogen
donor in reductive biosynthesis, and in defense
against oxidative stress. The HMP also serves
to convert G6P to ribose 5-phosphate (R5P),
which is required for nucleotide biosynthesis.
While the glycolytic pathway has been extensively
investigated in T. brucei, there have been
only a limited number of studies on HMP and
its contribution to carbohydrate metabolism.
The oxidative branch of the pathway converts
G6P to ribulose 5-phosphate (Ru5P), a process
which leads to the production of two moles of
NADPH per mole of G6P consumed. Recent
biochemical evidence, as well as sequence information,
suggests that several enzymes of this
BIOCHEMISTRY AND CELL BIOLOGY: PROTOZOA