21 Biochemistry of Fruits 499reactions in the form of NADPH (nicotinamide ade-
nine dinucleotide phosphate, reduced form), and sup-
plies carbon skeletons for the biosynthesis of several
secondary plant products. The organic acids stored in
the vacuole are metabolized through the functional
reversal of the respiratory pathway; this process is
termed gluconeogenesis. Altogether, sugar metabo-
lism is a key biochemical characteristic of the fruits.
In the glycolytic steps of reactions (Fig. 21.4),
glucose-6-phosphate is isomerized to fructose-6-
phosphate by the enzyme hexose phosphate isomer-
ase. Glucose-6-phosphate is derived from glucose-
1-phosphate by the action of glucose phosphate
mutase. Fructose-6-phosphate is phosphorylated at
the C1 position, yielding fructose-1,6-bisphosphate.
This reaction is catalyzed by the enzyme phospho-
fructokinase in the presence of ATP. Fructose-1,
6-bisphosphate is further cleaved into two, three-
carbon intermediates, dihydroxyacetone phosphate
and glyceraldehyde-3-phosphate, catalyzed by the
enzyme aldolase. These two compounds are inter-
convertible through an isomerization reaction medi-
ated by triose phosphate isomerase. Glyceraldehyde-
3-phosphate is subsequently phosphorylated at the
C1 position using orthophosphate, and oxidized
using NAD, to generate 1,3-diphosphoglycerate and
NADH. In the next reaction, 1,3-diphosphoglycerate
is dephosphorylated by glycerate-3-phosphate kin-
ase in the presence of ADP, along with the formation
of ATP. Glycerate-3-phosphate formed during this
reaction is further isomerized to 2-phosphoglycerate
in the presence of phosphoglycerate mutase. In the
presence of the enzyme enolase, 2-phosphoglycerate
is converted to phosphoenol pyruvate (PEP). De-
phosphorylation of PEP in the presence of ADP by
pyruvate kinase yields pyruvate and ATP. The meta-
bolic fate of pyruvate is highly regulated. Under nor-
mal conditions, it is converted to acetyl-CoA, which
then enters the citric acid cycle. Under anaerobic
conditions, pyruvate can be metabolized to ethanol,
which is a by-product in several ripening fruits.
There are two key regulatory steps in glycolysis,
one mediated by phosphofructokinase (PFK) and
the other by pyruvate kinase. In addition, there are
other types of modulation involving cofactors and
enzyme structural changes reported to be involved
in glycolytic control. ATP levels increase during
ripening. However, in fruits, this does not cause a
feedback inhibition of PFK like that observed in ani-
mal systems. There are two isozymes of PFK in
plants, one localized in plastids and the other in the
cytoplasm. These isozymes regulate the flow of car-
bon from the hexose phosphate pool to the pentose
phosphate pool. PFK isozymes are strongly inhibit-
ed by phosphoenol pyruvate (PEP). Thus, any con-
ditions that may cause the accumulation of PEP will
tend to reduce the carbon flow through glycolysis.
By contrast, inorganic phosphate is a strong activa-
tor of PFK. Thus, the ratio of PEP to inorganic phos-
phate would appear to be the major factor that regu-
lates the activity of PFK and carbon flux through
glycolysis. Structural alteration of PFK that increases
the efficiency of utilization of fructose-6-phosphate
is another means of regulation that can activate the
carbon flow through the glycolytic pathway.
Other enzymes of the glycolytic pathway are in-
volved in the regulation of starch/sucrose biosynthesis
(see Figs. 21.2 and 21.3). Fructose-1,6-bisphosphate
is converted back to fructose-6-phosphate by the
enzyme fructose-1,6-bisphosphatase, also releasing
inorganic phosphate. This enzyme is localized in the
cytosol and the chloroplast. Fructose-6-phosphate is
converted to fructose-2,6-bisphosphate by fructose-
6-phosphate 2-kinase, which can be dephosphoryla-
ted at the 2 position by fructose-2,6-bisphosphatase.
Fructose-6-phosphate is an intermediary in sucrose
biosynthesis (Fig. 21.3). Sucrose phosphate synthase
(SPS) is regulated by reversible phosphorylation (a
form of posttranslational modification that involves
addition of a phosphate moiety from ATP to an OH-
amino acid residue in the protein, such as serine or
threonine, mediated by a kinase; and dephosphory-
lation mediated by a phosphatase) by SPS kinase
and SPS phosphatase. Phosphorylation of the en-
zyme makes it less active. Glucose-6-phosphate is
an allosteric activator (a molecule that can bind to an
enzyme and increase its activity through enzyme
subunit association) of the active form of SPS (de-
phosphorylated). Glucose-6-phosphate is an inhib-
itor of SPS kinase, and inorganic phosphate is an in-
hibitor of SPS phosphatase. Thus, under conditions
when glucose-6-phosphate/inorganic phosphate ratio
is high, the active form of SPS will dominate, favor-
ing sucrose phosphate biosynthesis. These regula-
tions are highly complex and may be regulated by
the flux of other sugars in several pathways.
The conversion of PEP to pyruvate, mediated
by pyruvate kinase, is another key metabolic step
in the glycolytic pathway and is irreversible. Pyru-
vate is used in several metabolic reactions. During
respiration, pyruvate is further converted to acetyl
coenzyme A (acetyl-CoA), which enters the citric