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Photosynthesis
The carbon reactions
(Dark Reactions)
Overall Perspective
• Light reactions:
– Harvest light energy
– Convert light energy to
chemical energy
• Dark Reactions:
– Expend chemical energy
– Fix Carbon [convert CO2
to organic form]
At the end of the light reactions
• The reaction of the light reaction is:
– CO2 +H2O
(CH2O) + O2
• Recent estimates indicate that about 200 billion
tones of CO2 (Mr = 44) are converted to biomass
each year
– 40 % of this is from marine phytoplankton
– The bulk of the carbon is incorporated into organic
compounds by the carbon reducing reactions (dark
reactions) of photosynthesis
At the end of the light reactions
• The reactions catalyzing the
reduction of CO2 to
carbohydrates are coupled to the
consumption of NADPH and
ATP by enzymes found in the
stroma
– fluid environment
• These reactions were thought to
be independent of the light
reactions
– So the name “dark reactions” stuck
• However, these chemical
reactions are regulated by light
– So are called the “carbon reactions”
of photosynthesis
Overview of the carbon reactions
• The Calvin cycle:
• Stage 1:
– CO2 accepted by Ribulose-1,5bisphosphate.
– This undergoes carboxylation
• Has a carboxyl group
(-COOH) attached to it
– At the end of stage 1, CO2
covalently linked to a carbon
skeleton forming two
3-phosphycerate molecules.
Carboxylation: The first step is
the most important
• Step 1: The enzyme RUBISCO (Ribulose bis-phosphate carboxylase
oxygenase) carries out this conversion
• Rubisco accounts for 40% of the protein content of chloroplasts
– is likely the most abundant protein on Earth
• Rubisco is, in fact, very inefficient, and that a mechanism has evolved
to deal with this handicap
Overview of the carbon reactions
• The Calvin cycle:
• Stage 2:
– Each of the two 3-phosphycerate
molecules are altered.
– First phosphorylated through the
use of the 3 ATPs generated
during the light reaction.
– Then reduced through the use of
the 2 NADPHs generated during
the light reaction.
– Forms a carbohydrate
• glyceraldehyde-3-phosphate
3-phosphycerate molecules are
altered
• First phosphorylated through the use of the 3 ATP molecules
generated during the light reaction
– Forms 1,3-bisphosphoglycerate
• Then reduced through the use of the 2 NADPH molecules generated
during the light reaction
– Forms glyceraldehyde-3-phosphate
• Note the formation of triose phosphate
Overview of the carbon reactions
• The Calvin cycle:
• Stage 3:
– Regeneration of Ribulose-1,5bisphosphate.
– This requires the coordinated
action of eight reaction steps
• And thus eight specific enzymes
– Three molecules of Ribulose-1,5bisphosphate are formed from
the reshuffling of carbon atoms
from triose phosphate.
Regeneration of
Ribulose-1,5-bisphosphate
• The Calvin cycle reactions regenerate the
biochemical intermediates needed for operation
• More importantly, the cycle is Autocatalytic
– Rate of operation can be enhanced by increasing the
concentration of the intermediates in the cycle
• So, Calvin cycle has the metabolically desirable of
producing more substrate than is consumed
– Works as long as the produced triose phosphate is NOT
diverted elsewhere (as in times of stress or disease)
Overview of the carbon reactions
• The Calvin cycle:
• The cycle runs six times:
– Each time incorporating a new
carbon . Those six carbon
dioxides are reduced to glucose:
– Glucose can now serve as a
building block to make:
• polysaccharides
• other monosaccharides
• fats
• amino acids
• nucleotides
Only one-sixth of the triose phosphate
is used for polysaccharide production
• Synthesis of polysaccharides, such as starch and
sucrose, provide a sink
– Ensures an adequate flow of carbon atoms through the
cycle IF CO2 is constantly available
• During a steady rate of photosynthesis 5/6 of the
triose phosphates are used for the regeneration of
Ribulose-1,5-bisphosphate
• 1/6 is transported to the cytosol for the synthesis of
sucrose or other metabolites that are converted to
starch in the chloroplast
Regulation of the Calvin cycle
• The high energy efficiency of the Calvin cycle
indicates that some form of regulation ensures that
all intermediates in cycle:
– Are present at adequate concentrations
– The cycle is turned off when it is not needed in the dark
• Remember:
– These are the “carbon reactions”, NOT the “dark reactions”
• Many factors regulate the Calvin cycle
Regulation of the Calvin cycle
• 1: The pH of the stroma increases as protons are
pumped out of it through the membrane assembly of
the light reactions.
– The enzymes of the Calvin Cycle function better at this
higher pH.
• 2: The reactions of the Calvin cycle have to stop
when they run out of substrate
– as photosynthesis stops, there is no more ATP or
NADPH in the stroma for the dark reactions to take
place.
Regulation of the Calvin cycle
• 3: The light reactions increase the permeability of
the stromal membrane to required cofactors
– Mg ions are required for the Calvin Cycle.
• 4: Several enzymes of the Calvin Cycle are
activated by the breaking of disulphide bridges of
enzymes involved in the working of the cycle.
– the activity of the light reactions is communicated to the
dark reactions by an enzyme intermediate
When conditions are
not optimum
Photorespiration
• Occurs when the CO2 levels inside a leaf become
low
– This happens on hot dry days when a plant is forced to
close its stomata to prevent excess water loss
• If the plant continues to attempt to fix CO2 when its
stomata are closed
– CO2 will get used up and the O2 ratio in the leaf will
increase relative to CO2 concentrations
• When the CO2 levels inside the leaf drop to around
50 ppm,
– Rubisco starts to combine O2 with Ribulose-1,5bisphosphate instead of CO2
Photorespiration
• Instead of producing 2 3C PGA
molecules, only one molecule of
PGA is produced and a toxic 2C
molecule called phosphoglycolate
is produced
• The plant must get rid of the
phosphoglycolate
• The plant immediately gets rid of
the phosphate group
– converting the molecule to
glycolic acid
Photorespiration
• The glycolic acid is then
transported to the peroxisome and
there converted to glycine
– Peroxisomes are ubiquitous
organelles that function to rid
cells of toxic substances
• The glycine (4 carbons) is then
transported into a mitochondria
where it is converted into serine
(3 carbons)
– Releases CO2
Photorespiration
• The serine is then used to make
other organic molecules
• All these conversions cost the
plant energy and results in the net
lost of CO2 from the plant
• 75% of the carbon lost during the
oxygenation of Rubisco is
recovered during photorespiration
and is returned to the Calvin cycle
The C4 Carbon
cycle
The C4 carbon Cycle
• The C4 carbon Cycle occurs in 16 families of both
monocots and dicots.
–
–
–
–
Corn
Millet
Sugarcane
Maize
• There are three variations of the basic C4 carbon
Cycle
– Due to the different four carbon molecule used
The C4 carbon Cycle
• This is a biochemical pathway
that prevents photorespiration
• C4 leaves have TWO chloroplast
containing cells
– Mesophyll cells
– Bundle sheath (deep in the leaf so
atmospheric oxygen cannot diffuse easily to
them)
• C3 plants only have Mesophyll cells
• Operation of the C4 cycle requires the
coordinated effort of both cell types
– No mesophyll cells is more than
three cells away from a bundle
sheath cells
• Many plasmodesmata for
communication
The C4 carbon Cycle
• Four stages:
• Stage 1:
• In Mesophyll cell
– Fixation of CO2 by the carboxylation
of phosphenol-pyruvate (primary
acceptor molecule)
– forms a C4 acid molecule
– Malic acid and/or aspartate
• Stage 2:
– Transport of the C4 acid molecule to
the bundle sheath cell
The C4 carbon Cycle
• Stage 3:
– Decarboxylation of the C4 acid
molecule (in bundle sheath)
– Makes a C3 acid molecule
– This generates CO2
– This CO2 is reduced to carbohydrate
by the Calvin cycle
• Stage 4:
– The C3 acid molecule (pryuvate) is
transported back to mesophyll cells
– Regeneration of phosphenolpyruvate
The C4 carbon Cycle
• Regeneration of phosphenol-pyruvate consumes two high energy
bonds from ATP
• Movement between cells is by diffusion via plasmodesmata
• Movement within cells is regulated by concentration gradients
• This system generates a higher CO2 conc in bundle sheath cells than
would occur by equilibrium with the atmosphere
– Prevents photorespiration!!!!!!!!!!
The C4 carbon Cycle
• The net effect of the C4 carbon Cycle is to convert a dilute
CO2 solution in the mesophyll into a concentrated solution
in the bundle sheath cells
– This requires more energy than C3 carbon plants
• BUT – This energy requirement is constant no matter what
the environmental conditions
• Allows more efficient photosynthesis in hotter conditions
Crassulacean Acid
Metabolism
(CAM Plants)
CAM Plants
• The CAM mechanism enables plants to improve
water efficiency
– CAM plant
• Loses 50 – 100 g water for every gram of CO2 gained
– C4 plant
• Loses 250 – 300 g water for every gram of CO2 gained
– C3 plant
• Loses 400 – 500 g water for every gram of CO2 gained
• Similar to C4 cycle
– In CAM plants formation of the C4 acid is both
temporally and spatially separated
CAM Plants
• At night:
• Stomata only open at night when
it is cool
• CO2 is captured by phosphenolpyruvate carboxylase in the
cytosol – leaves become acidic
• The malic acid formed is stored
in the vacuole
– Amount of malic acid formed is
equal to the amount of CO2 taken
in
CAM Plants
• During the day:
• Stomata close, preventing water loss,
and further uptake of CO2
• Malic acid is transported to the
chloroplast and decarboxylated to
release CO2
• This enters the Calvin cycle as it can
not escape the leaf
– Pyruvate is converted to starch in the
chloroplast – regenerates carbon
acceptor
Phosphorylation regulates phosphenolpyruvate (PEP) carboxylase
• CAM and C4 plants require a
separation of the initial
carboxylation from the following
de-carboxylation
• Diuranal regulation is used
• IN CAM PLANTS:• Phosphorylation of the serine residue of
phosphenol-pyruvate (PEP) carboxylase
(Ser-OP) yields a form of the enzyme
which is active at night
– This is relatively insensitive to malic
acid
Photophorylation regulates phosphenolpyruvate (PEP)carboxylase
• During the day:
• De-Phosphorylation of the serine
(ser-OH) gives a form of the
enzyme which is inhibited by malic
acid
• THIS IS THE OPPOSITE WAY
AROUND FOR C4 PLANTS!
Summary
• The reduction of CO2 to carbohydrate via
photosynthesis is coupled to the consumption of
ATP and NADPH
• CO2 is reduced via the Calvin cycle
– Takes place in the stroma (soluble phase)
• CO2 and water combine with Ribulose-1,5bisphosphate in the following reaction
– CO2 +H2O
(CH2O) + O2
• Regeneration of the carrier is required for the cycle
to continue
Summary
• The Calvin cycle requires the joint action of several
light-dependant systems
– Changes in ions (Mg+ and H+)
– Changes in effector metabolites (enzyme substrates)
– Changes in protein-mediated systems (rubisco activase)
• Rubisco can also act as an oxygenase
– The carboxylation & oxygenation reactions take place
at the active sites of rubisco.
Summary
• C4 and CAM plants Prevent photorespiration!!!!!
• C4 leaves have TWO chloroplast containing cells
– Mesophyll cells
– Bundle sheath
• CAM Plants drastically reduce water lass
– CAM plant
• Loses 50 – 100 g water for every gram of CO2 gained
– C4 plant
• Loses 250 – 300 g water for every gram of CO2 gained
– C3 plant
• Loses 400 – 500 g water for every gram of CO2 gained
Any Questions?