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C-4 PHOTOSYNTHESIS: INVOLVES “ADD-ON” THAT ALLOWS SOME
PLANTS TO USE CO2 MORE EFFECTIVELY
C-3 photosynthesis evolved first:
1) Photosynthesis evolved (in cyanobacteria) when the [CO2] was much higher than it
is now and when the [O2] was much lower.
2) Hence there was no selective pressure for an efficient CO2-fixing enzyme to evolve.
And one didn’t!
3) The enzyme that evolved was RuBisCO (ribulose bis-phosphate carboxylase
oygenase). It is clear why a CO2-fixing enzyme would be called a carboxylase. It is
not so obvious at all why it would also be called an oxygenase (an enzyme that
“fixes”, or incorporates O2 molecules into organic compounds. More about that
later.
4) RuBisCO catalyzes the following reaction:
RuBP
5C
+
CO2
2 molecules PGA
1C
RuBP = ribulose-bis-phosphate
2 x 3C
PGA = phosphoglyceric acid
5) RuBisCO is a very bad enzyme (catalyst)!
a) It has a low affinity for CO2 (does not bind it into its active site efficiently).
In fact, at present day [CO2] levels it is only working at about 70% of its
maximal velocity (Vmax).
b) Even when working at its Vmax (active site is saturated with CO2),
RuBisCO is a very slow enzyme. The maximum number of times that a
single RuBisCO molecule can perform a reaction (its so-called turnover
number) is 4 times a second (4s-1)! This is a terrible!
c) RuBisCO does not distinguish between CO2 and O2 as well as it should.
Because the atmospheric [CO2] only allows RuBisCO to work at about 70%
of its Vmax (a), because its Vmax is equivalent to only 4 reactions per second
(b), and because (c) O2 can displace CO2 in the active site, and take part in a
reaction (the oxygenase reaction, see figure below) it turns out RuBisCO can
only catalyze, on average, 2 actual carboxylase reactions a minute (2s-1)!
2
5C
1C
RuBP CO2
3C
3C
PGA PGA
RuBisCO
High [CO2] and low [O2] conditions.
THIS IS THE CARBOXYLASE REACTION
5C
RuBP
3C
2C
PGA P-Glycolate
0C
O2
RuBisCO
Normal atmospheric [CO2] and [O2]
conditions about 1 out 3 reactions is
the above.
THIS IS THE OXYGENASE REACTION
A plant out in the field will be photosynthesizing under the conditions described below:
1) The atmospheric [CO2] is low so that each RuBisCO molecule is only operating at about
3s-1 (i.e. about 70% of Vmax = 4s-1).
2) The atmospheric [O2] is so high that in about 1 out 3 reactions, an O2 molecule outcompetes a CO2 molecule for entry into the RuBisCO active site and actually takes part
in a the oxygenase reaction. In this case, RuBisCO catalyses the addition of the O2
molecule to RuBP and not the addition of a CO2 molecule (the carboxylase function of
RuBisCO). Hence, under atmospheric conditions the actual average turnover number for
a RuBisCO molecule is only about 2s-1!
3) So RuBisCO can be a carboxylase (the reaction shown on the left in the above diagram),
and do what we expect it to do in photosynthesis and “fix” CO2 into organic molecules.
Or (because it does not distinguish O2 from CO2 as well as it should) it can act as an
oxygenase and “fix” CO2 into organic molecules. In terms of “fixing” CO2, the
oxygenase reaction (shown on the right in the above diagram) is a wasted reaction.
What is a C-3 photosynthesis?
This is the type of photosynthesis you learned about in previous lectures. It is by far the
most common type of photosynthesis occurring in Nova Scotia! It is called C-3 photosynthesis because
its first product is a 3C-containing compound called 3-phosphosphoglyceric acid (or 3phosphoglycerate) i.e. the PGA in the above diagrams. Plants that only do this type of photosynthesis
are called C-3 plants.
What is a C-4 photosynthesis?
In this type of photosynthesis the first compound made is not PGA but malic acid
(malate) which has 4C. Plants that have this type of photosynthesis are called C-4 plants. Many of the C4 plants are grasses that first arose in rather dry areas in the tropics. This type of photosynthesis was
first discovered in sugar cane. Another well known example is maize (corn) (Zea mays).
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Open stomata let in CO2 but allow H2O to escape.
Stomata have been called a “necessary evil” in terms of the physiology of plants. You
will already have learned of the various morphological tricks that plants use to reduce water loss from
their stomata (stomates). These include location of the greatest number of stomates on the shaded
bottom of a leave than the top and the sunken stomates of conifer needles. C-4 photosynthesis is a
biochemical (with related morphology) trick to reduce water loss from the leaves while still being able
to fix CO2 effectively. It is not surprising that C-4 photosynthesis evolved in the tropics (warm) in dry
places. One finds C-4 plants on the African plains, for example, that as one climbs up Mount
Kilimanjaro and it gets cooler and more moist the C-3 plants become more abundant. In Nova Scotia our
native plants that have C-4 metabolism are mainly grasses growing in the salt marshes where roots have
trouble obtaining water because of the high osmotic potential of seawater. In other words, these slat
marsh plants have to conserve water as much as if they were growing in a dry place.
All grasses tend to roll up their rims (whoops-blades) during the middle of the day when
it is hottest. But when C-3 plants do this their rate of photosynthesis slows down greatly and less CO2
can enter the stomata of the rolled up leaf, and the stomates themselves also at least partially close.
Water loss is slowed down but so is CO2 entry, and we know how poor C-3 plants are at fixing CO2 at
the best of times, dependent as they are on the inefficient RuBisCO! What if a plant had a better enzyme
to do the fixation rather than RuBisCO? On this the following comments:
1) RuBisCO is a very inefficient enzyme. But it does seem as if selective pressures
have at least made it as good as it can be (which is lousy)! So it does not seem as
if RuBisCO can be improved.
2) The rest of the Calvin Cycle does it job quite well with “competent” enzymes.
3) So what has evolved in C-4 plants is not a completely different way of making the
photosynthetic intermediates but, instead, an “add-on” that ultimately makes use
of the Calvin Cycle but provides conditions under which RuBisCO can at least
operate at its Vmax (4s-1!). In other words, the “add-on” of C-4 photosynthesis
allows the rate of CO2 –fixation to be about twice as high as in C-3 plants (only
2s-1). Actually, even more importantly, the C-4 “add-on” allows the plants to
operate efficiently at very low [CO2]. This is what is needed if a sugar cane or
corn plant is to be able to perform adequate photosynthesis when only low rates of
CO2 uptake are occurring through partially closed stomates hidden in a rolled up
leaf.
4) The “better enzyme” in C-4 photosynthesis is PEP carboxylase. (PEP is a 3Ccontaining compound, phosphoenolpyruvate. (Those of you taking Biochemistry
should know about is role in glycolysis.). PEP carboxylase is everything that
RuBisCO isn’t. It has a high affinity for its substrate (bicarbonate, not CO2), it
turnover (operates) many times a second, and it has no affinity for O2. (The latter
is not surprising since as a molecule O2 has no structural similarity to bicarbonate,
HCO3-1. Any active site evolved to bind HCO3-1 will not be able to bind O2
instead by accident. However, O2 does have structural similarities to CO2, so the
active site of RuBisCO can accept O2 by accident, especially since the
atmospheric [O2] is much greater than the [CO2].
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5) PEP carboxylase is found in the mesophyll cells of C-4 plants.
6) Unlike C-3 plants, whose bundle sheath cells are dead, the bundle sheath cells are
big cells with big chloroplasts and a very alive. This is where the RUBisCO of C4 plants is located. There is no RuBisCO in the mesophyll cells and no PEP
carboxylase in the bundle sheath cells. (In C-3 plants the RuBisCO is in the
mesophyll cells, spongy and palisade, and PEP carboxylase is found only in low
amounts and is not directly involved in photosynthesis.)
7)
This diagram outlines what is going-on in C4 photosynthesis:
1) CO2 enters, slowly through the partially
closed stomata of the rolled-up leaf.
2) Most of the CO2 is rapidly converted to
bicarbonate, HCO3-, by an enzyme called
carbonic anhydrase. (Unfortunately this
diagram does not show this important step!)
This enzyme is probably the fastest enzyme
in the world, unlike RuBisCO which is probably the slowest enzyme in the world, and each
molecule of this carbonic anhydrase can catalyze almost 106 reactions a second!
3) The HCO3- is then joined to PEP in the active site of PEP carboxylase. (Enzymes are called
carboxlyases whether it is CO2 or HCO3-1 that they use as a substrate. There is no
carboxylase known that can use both CO2 and HCO3-1, they use one or the other always. An
active site that could use either CO2 or HCO3-1 is not possible, since the molecule and the ion
are so structurally different.
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4) The product of the PEP carboxylase reaction is called OAA, but this is very rapidly
converted to malate (malic acid). No need to remember the OAA, but do remember malate.
Both OAA and malate have the same number of C-atoms i.e 4.
PEP + HCO3-1
3C
1C
OAA
4C
Malate
4C
(don’t worry about OAA)
PEP carboxylase
5) The malate then diffuses, through plasmodesmata, from one mesophyll cell to another until it
reaches a bundle sheath cell.
6) Once in a bundle sheath cell, the malate is decarboxlyated and CO2 (not HCO3-1) is released
into the bundle sheath cell. The CO2 diffuses into the large chloroplast, and the Calvin Cycle,
including RuBisCO takes over!
7) What the malate molecules are doing is carrying (in the form of an attached carboxyl group)
the CO2 molecules, that “dribbled” into the rolled-up leaf, from the mesophyll cells to the
bundle sheath cells. In the bundle sheath cells the malate molecules are decarboxylated (by
an enzyme) and the CO2 is used by RuBisCO in the normal way. But the [CO2] in the bundle
sheath becomes quite high, due to the decarboxylation of all the malate that diffuses into the
bundle sheath cells, and so the RuBisCO works at its Vmax.
Malate
4C
Pyruvate + CO2
3C
1C
CO2 + RuBP
2 molecules PGA
8) The pyruvate goes back to the mesophyll cells and is converted to PEP.
Pyruvate
PEP
3C
3C
ADP
ADP
This conversion requires ATP (pyruvate contains no phosphate, but
phospoenolpyruvate does!). This use of ATP is part of the extra energy that is needed to drive
C-4 photosynthesis compared to C-3 photosynthesis.
9) So the [CO2] in the bundle sheath cells helps the RuBisCO to work better. What also helps is
that the chloroplast of the bundle sheath cells are quite different from those of the mesophyll
cells. Unlike most chloroplasts the chloroplasts in the bundle sheath cells do not have
photosystem II. So they do not produce O2! So the “oxygenase problem” of RuBisCO is also
solved in the bundle sheath cells!
Material in blue font was not mentioned in the lecture.
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RuBisCO
Ribulose bisphosphate carboxylase oxygenase
RuBisCO is also written as RUBISCO. (slow enzyme, big name!)