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Carbohydrate
Biosynthesis in Plants
and Bacteria
chapter
20
S-238
1. Segregation of Metabolism in Organelles What are the advantages to the plant cell of having
different organelles to carry out different reaction sequences that share intermediates?
2. Phases of Photosynthesis When a suspension of green algae is illuminated in the absence of CO
2
and then incubated with
14
CO
2
in the dark,
14
CO
2
is converted to [
14
C]glucose for a brief time. What is
the significance of this observation with regard to the CO
2
-assimilation process, and how is it related to
the light reactions of photosynthesis? Why does the conversion of
14
CO
2
to [
14
C]glucose stop after a brief
time?
3. Identification of Key Intermediates in CO
2
Assimilation Calvin and his colleagues used the
unicellular green alga Chlorella to study the carbon-assimilation reactions of photosynthesis. They
incubated
14
CO
2
with illuminated suspensions of algae and followed the time course of appearance of
14
C in two compounds, X and Y, under two sets of conditions. Suggest the identities of X and Y, based
on your understanding of the Calvin cycle.
(a) Illuminated Chlorella were grown with unlabeled CO
2
, then the light was turned off and
14
CO
2
was added (vertical dashed line in graph (a)). Under these conditions, X was the first compound
to become labeled with
14
C; Y was unlabeled.
(b) Illuminated Chlorella cells were grown with
14
CO
2
. Illumination was continued until all the
14
CO
2
had disappeared (vertical dashed line in graph (b)). Under these conditions, X became labeled
quickly but lost its radioactivity with time, whereas Y became more radioactive with time.
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4. Regulation of the Calvin Cycle Iodoacetate reacts irreversibly with the free OSH groups of Cys
residues in proteins.
Chapter 20 Carbohydrate Biosynthesis in Plants and Bacteria S-239
Radioactivity
CO2,
light
0
Time
14CO2,
dark
Radioactivity
0
Time
14CO2
Y
X
Y
X
(a)
(b)
C
OHI
O
Iodoacetate Inactive enzyme
NAD
SH
Cys ICH
2
C
O
O
CH
2
NAD
S
Cys
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S-240 Chapter 20 Carbohydrate Biosynthesis in Plants and Bacteria
Predict which Calvin cycle enzyme(s) would be inhibited by iodoacetate, and explain why.
5. Thioredoxin in Regulation of Calvin Cycle Enzymes Motohashi and colleagues used thioredoxin
as a hook to fish out from plant extracts the proteins that are activated by thioredoxin. To do this,
they prepared a mutant thioredoxin in which one of the reactive Cys residues was replaced with a Ser.
Explain why this modification was necessary for their experiments. [Motohashi, K., Kondoh, A.,
Stumpp, M.T., & Hisabori, T. (2001) Comprehensive survey of proteins targeted by chloroplast thiore-
doxin. Proc. Natl. Acad. Sci. USA 98, 11,224-11,229.]
6. Comparison of the Reductive and Oxidative Pentose Phosphate Pathways The reductive
pentose phosphate pathway generates a number of intermediates identical to those of the oxidative
pentose phosphate pathway (Chapter 14). What role does each pathway play in cells where it is active?
7. Photorespiration and Mitochondrial Respiration Compare the oxidative photosynthetic carbon
cycle (C
2
cycle), also called photorespiration, with the mitochondrial respiration that drives ATP
synthesis. Why are both processes referred to as respiration? Where in the cell do they occur, and un-
der what circumstances? What is the path of electron flow in each?
8. Rubisco and the Composition of the Atmosphere N. E. Tolbert has argued that the dual speci-
ficity of rubisco for CO
2
and O
2
is not simply a leftover from evolution in a low-oxygen environment.
He suggests that the relative activities of the carboxylase and oxygenase activities of rubisco actually
have set, and now maintain, the ratio of CO
2
to O
2
in the earth’s atmosphere. Discuss the pros and
cons of this hypothesis, in molecular terms and in global terms. How does the existence of C
4
organ-
isms bear on the hypothesis? [Tolbert, N.E. (1994) The role of photosynthesis and photorespiration in
regulating atmospheric CO
2
and O
2
. In Regulation of Atmospheric CO
2
and O
2
by Photosynthetic
Carbon Metabolism (Tolbert, N.E. & Preiss, J., eds), pp. 8–33, Oxford University Press, New York.]
9. Role of Sedoheptulose 1,7-Bisphosphatase What effect on the cell and the organism might result from
a defect in sedoheptulose 1,7-bisphosphatase in (a) a human hepatocyte and (b) the leaf cell of a green
plant?
Answer
10. Pathway of CO
2
Assimilation in Maize If a maize (corn) plant is illuminated in the presence of
14
CO
2
, after about 1 second, more than 90% of all the radioactivity incorporated in the leaves is found
at C-4 of malate, aspartate, and oxaloacetate. Only after 60 seconds does
14
C appear at C-1 of
11. Identifying CAM Plants Given some
14
CO
2
and all the tools typically present in a biochemistry re-
search lab, how would you design a simple experiment to determine whether a plant was a typical C
4
plant or a CAM plant?
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S-242 Chapter 20 Carbohydrate Biosynthesis in Plants and Bacteria
12. Chemistry of Malic Enzyme: Variation on a Theme Malic enzyme, found in the bundle-sheath
cells of C
4
plants, carries out a reaction that has a counterpart in the citric acid cycle. What is the
analogous reaction? Explain your choice.
13. The Cost of Storing Glucose as Starch Write the sequence of steps and the net reaction required
to calculate the cost, in ATP molecules, of converting a molecule of cytosolic glucose 6-phosphate to
starch and back to glucose 6-phosphate. What fraction of the maximum number of ATP molecules
available from complete catabolism of glucose 6-phosphate to CO
2
and H
2
O does this cost represent?
14. Inorganic Pyrophosphatase The enzyme inorganic pyrophosphatase contributes to making many
biosynthetic reactions that generate inorganic pyrophosphate essentially irreversible in cells. By keep-
ing the concentration of PP
i
very low, the enzyme “pulls” these reactions in the direction of PP
i
formation. The synthesis of ADP-glucose in chloroplasts is one reaction that is pulled in the forward
direction by this mechanism. However, the synthesis of UDP-glucose in the plant cytosol, which
produces PP
i
, is readily reversible in vivo. How do you reconcile these two facts?
15. Regulation of Starch and Sucrose Synthesis Sucrose synthesis occurs in the cytosol and starch
synthesis in the chloroplast stroma, yet the two processes are intricately balanced. What factors shift
the reactions in favor of (a) starch synthesis and (b) sucrose synthesis?
Answer
16. Regulation of Sucrose Synthesis In the regulation of sucrose synthesis from the triose phosphates
produced during photosynthesis, 3-phosphoglycerate and P
i
play critical roles (see Fig. 20–25).
Explain why the concentrations of these two regulators reflect the rate of photosynthesis.
17. Sucrose and Dental Caries The most prevalent infection in humans worldwide is dental caries,
which stems from the colonization and destruction of tooth enamel by a variety of acidifying
microorganisms. These organisms synthesize and live within a water-insoluble network of dextrans,
called dental plaque, composed of (a1 n6)-linked polymers of glucose with many (a1 n3) branch
points. Polymerization of dextran requires dietary sucrose, and the reaction is catalyzed by a bacterial
enzyme, dextran-sucrose glucosyltransferase.
(a) Write the overall reaction for dextran polymerization.
(b) In addition to providing a substrate for the formation of dental plaque, how does dietary sucrose
also provide oral bacteria with an abundant source of metabolic energy?
18. Differences between C
3
and C
4
Plants The plant genus Atriplex includes some C
3
and some C
4
species. From the data in the plots below (species 1, upper curve; species 2, lower curve), identify which
is a C
3
plant and which is a C
4
plant. Justify your answer in molecular terms that account for the data in
all three plots.
Chapter 20 Carbohydrate Biosynthesis in Plants and Bacteria S-243
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S-244 Chapter 20 Carbohydrate Biosynthesis in Plants and Bacteria
19. C
4
Pathway in a Single Cell In typical C
4
plants, the initial capture of CO
2
occurs in one cell type,
and the Calvin cycle reactions occur in another (see Fig. 20–23). Voznesenskaya and colleagues have
described a plant, Bienertia cycloptera—which grows in salty depressions of semidesert in Central
Asia—that shows the biochemical properties of a C
4
plant but unlike typical C
4
plants does not segre-
gate the reactions of CO
2
fixation into two cell types. PEP carboxylase and rubisco are present in the
same cell. However, the cells have two types of chloroplasts, which are localized differently, as shown
in the micrograph. One type, relatively poor in grana (thylakoids), is confined to the periphery; the
more typical chloroplasts are clustered in the center of the cell, separated from the peripheral chloro-
plasts by large vacuoles. Thin cytosolic bridges pass through the vacuoles, connecting the peripheral
and central cytosol. [Voznesenskaya, E.V., Fraceschi, V.R., Kiirats, O., Artyusheva, E.G., Freitag, H., &
Edwards, G.E. (2002) Proof of C
4
photosynthesis without Kranz anatomy in Bienertia cycloptera
(Chenopodiaceae). Plant J. 31, 649–662.]
Uptake of CO2
Light intensity Leaf temperature [CO2] in intracellular space
10 m
In this plant, where would you expect to find (a) PEP carboxylase, (b) rubisco, and (c) starch gran-
ules? Explain your answers with a model for CO
2
fixation in these C
4
cells.
Data Analysis Problem
20. Rubisco of Bacterial Endosymbionts of Hydrothermal Vent Animals Undersea hydrothermal
vents support remarkable ecosystems. At these extreme depths there is no light to support photosyn-
thesis, yet thriving vent communities are found. Much of their primary productivity occurs through
chemosynthesis carried out by bacterial symbionts that live in specialized organs (trophosomes) of
certain vent animals.
Chemosynthesis in these bacteria involves a process that is virtually identical to photosynthesis.
Carbon dioxide is fixed by rubisco and reduced to glucose, and the necessary ATP and NADPH are
produced by electron-transfer processes similar to those of the light-dependent reactions of photosyn-
thesis. The key difference is that in chemosynthesis, the energy driving electron transfer comes from a
highly exergonic chemical reaction rather than from light. Different chemosynthetic bacteria use dif-
ferent reactions for this purpose. The bacteria found in hydrothermal vent animals typically use the
oxidation of H
2
S (abundant in the vent water) by O
2
, producing elemental sulfur. These bacteria also
use the conversion of H
2
S to sulfur as a source of electrons for chemosynthetic CO
2
reduction.
(a) What is the overall reaction for chemosynthesis in these bacteria? You do not need to write a
balanced equation; just give the starting materials and products.
(b) Ultimately, these endosymbiotic bacteria obtain their energy from sunlight. Explain how this
occurs.
Robinson and colleagues (2003) explored the properties of rubisco from the bacterial endosym-
biont of the giant tube worm Riftia pachyptila. Rubisco, from any source, catalyzes the reaction of
either CO
2
(Fig. 20–7) or O
2
(Fig. 20–20) with ribulose 1,5-bisphosphate. In general, rubisco reacts
more readily with CO
2
than O
2
. The degree of selectivity () can be expressed as
where Vis the reaction velocity.
Robinson and coworkers measured the value for the rubisco of the bacterial endosymbionts.
They purified rubisco from tube-worm trophosomes, reacted it with mixtures of different ratios of O
2
and CO
2
in the presence of [1-
3
H]ribulose 1,5-bisphosphate, and measured the ratio of [
3
H]phospho-
glycerate to [
3
H]phosphoglycolate.
(c) The measured ratio of [
3
H]phosphoglycerate to [
3
H]phosphoglycolate is equal to the ratio
V
carboxylation
兾V
oxygenation
. Explain why.
(d) Why would [5-
3
H]ribulose 1,5-bisphosphate not be a suitable substrate for this assay?
The for the endosymbiont rubisco had a value of 8.6 0.9.
(e) The atmospheric (molar) concentration of O
2
is 20% and that of CO
2
is about 380 parts per million. If
the endosymbiont were to carry out chemosynthesis under these atmospheric conditions, what
would be the value of V
carboxylation
/V
oxygenation
?
(f) Based on your answer to (e), would you expect for the rubisco of a terrestrial plant to be
higher than, the same as, or lower than 8.6? Explain your reasoning.
Two stable isotopes of carbon are commonly found in the environment: the more abundant
12
C
and the rare
13
C. All rubisco enzymes catalyze the fixation of
12
CO
2
faster than that of
13
CO
2
. As a
result, the carbon in glucose is slightly enriched in
12
C compared with the isotopic composition of CO
2
in the environment. Several factors are involved in this “preferential” use of
12
CO
2
, but one factor is
the fundamental physics of gases. The temperature of a gas is related to the kinetic energy of its mole-
cules. Kinetic energy is given by 1⁄2mv
2
, where mis molecular mass and vis velocity. Thus, at the same
temperature (same kinetic energy), the molecules of a lighter gas will be moving faster than those of a
heavier gas.
(g) How could this contribute to rubisco’s “preference” for
12
CO
2
over
13
CO
2
?
Some of the first convincing evidence that the tube-worm hosts were obtaining their fixed carbon
from the endosymbionts was that the
13
C/
12
C ratio in the animals was much closer to that of the bacte-
ria than that of nonvent marine animals.
[CO
2
]
[O
2
]
V
carboxylation
V
oxygenation
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S-246 Chapter 20 Carbohydrate Biosynthesis in Plants and Bacteria
(h) Why is this more convincing evidence for a symbiotic relationship than earlier studies that simply
showed the presence of rubisco in the bacteria found in trophosomes?
Answer
