C. Metabolic Processes · Metabolic ProcessesThe Calvin Cycle, Photorespiration, and C₄ and CAM Plants
C3.2 C2.1 C3.3 C1.1 A2.2About 70 minutes
Learning goals- I can describe the three phases of the Calvin cycle: carbon fixation, reduction and regeneration of RuBP.
- I can calculate the ATP, NADPH and CO₂ needed to make one G3P and one glucose.
- I can explain photorespiration and why it reduces the efficiency of photosynthesis.
- I can compare C₃, C₄ and CAM plants and relate their adaptations to their environments.
Words to know
carbon fixation
Incorporating CO₂ from the air into an organic molecule; in the Calvin cycle, CO₂ is attached to RuBP by rubisco.
rubisco
Ribulose bisphosphate carboxylase/oxygenase, the enzyme that fixes CO₂ to RuBP; probably the most abundant protein on Earth.
RuBP (ribulose 1,5-bisphosphate)
The five-carbon sugar that accepts CO₂ in the first step of the Calvin cycle and is regenerated at the end.
3-PGA (3-phosphoglycerate)
The three-carbon molecule formed when the unstable six-carbon product of CO₂ fixation splits.
G3P (glyceraldehyde 3-phosphate)
The three-carbon sugar produced by the Calvin cycle; used to make glucose, sucrose, starch and other molecules.
photorespiration
A wasteful process in which rubisco binds O₂ instead of CO₂, releasing CO₂ and using ATP without making sugar; worst in hot, dry conditions.
C₄ plant
A plant that first fixes CO₂ into a four-carbon acid in mesophyll cells, then releases it to the Calvin cycle in bundle-sheath cells (e.g. corn, sugar cane).
CAM plant
A plant that opens its stomata at night to fix CO₂ into acids, then uses the stored CO₂ in the Calvin cycle during the day with stomata closed (e.g. pineapple, cacti, jade plant).
PEP carboxylase
The enzyme that fixes CO₂ into a four-carbon acid in C₄ and CAM plants; unlike rubisco, it does not bind O₂.
Building sugar out of thin air
A large sugar maple can weigh several tonnes, and almost all of its dry mass is carbon, hydrogen and oxygen. The carbon came from the air, about 0.04% of which is CO₂. Grabbing that scarce CO₂ and turning it into sugar is the job of the Calvin cycle. In this lesson, you'll follow carbon atoms through the cycle, count the ATP and NADPH it costs, and see how plants such as Ontario's corn deal with the cycle's biggest weakness.
Remember from Lessons 4, 11 and 12
From Lessons 11 and 12: the Calvin cycle takes place in the stroma. It uses the ATP and NADPH made by the light reactions and returns ADP, Pᵢ and NADP⁺. From Lesson 4: G3P is also an intermediate of glycolysis.
Discovering the cycle
In the late 1940s and 1950s, Melvin Calvin, Andrew Benson and James Bassham at the University of California, Berkeley, fed radioactive ¹⁴CO₂ to the green alga Chlorella for a few seconds, then quickly killed the cells in hot alcohol and separated the compounds by paper chromatography. After the shortest exposures, the radioactive carbon showed up first in 3-PGA; with longer exposures, it appeared in sugars and then in RuBP. By working out the order in which compounds became labelled, they mapped the cycle. Calvin received the 1961 Nobel Prize in Chemistry. This is a great example of using isotopes and chromatography (the same technique you used on spinach pigments) to answer a biological question.
Phase 1: carbon fixation
The enzyme rubisco attaches CO₂ to RuBP, a five-carbon sugar. The six-carbon product is unstable and immediately splits into two molecules of 3-PGA (3C each).
3 CO₂ + 3 RuBP (15 C) → 6 3-PGA (18 C)
Rubisco is slow: each molecule fixes only a few CO₂ per second. To compensate, leaves contain huge amounts of it, and it is probably the most abundant protein on Earth.
Phase 2: reduction
Each 3-PGA is phosphorylated by ATP and then reduced by NADPH to form G3P. This is where the energy from the light reactions is stored in the sugar.
6 3-PGA + 6 ATP + 6 NADPH → 6 G3P
Phase 3: regeneration of RuBP
For every 3 CO₂ fixed, there are 6 G3P (18 C). One G3P (3 C) leaves the cycle as the net product. The other five G3P (15 C) are rearranged, using 3 ATP, into 3 RuBP (15 C), ready to fix more CO₂.
5 G3P + 3 ATP → 3 RuBP
The bill for one glucose
- Per G3P (3 CO₂): 9 ATP and 6 NADPH.
- Per glucose (6 CO₂, made from 2 G3P): 18 ATP and 12 NADPH.
Notice the ratio: 3 ATP for every 2 NADPH, which is why chloroplasts need cyclic electron flow to make extra ATP (Lesson 12).
What happens to G3P? It is the starting point for almost all organic molecules in a plant:
- two G3P → glucose (and fructose), which can be used in the plant's own cellular respiration (plant cells have mitochondria too!);
- glucose + fructose → sucrose, transported in the phloem to roots, fruits and growing tips;
- glucose → starch (storage, in chloroplasts and roots) and cellulose (cell walls);
- G3P carbon skeletons are also used to make amino acids and lipids.
Rubisco's flaw: photorespiration
Rubisco evolved more than two billion years ago, when the atmosphere had very little O₂. It can bind O₂ instead of CO₂ (that is the "oxygenase" in its name). When it does, RuBP is split into one 3-PGA and a two-carbon compound that the plant must recycle, a process that uses ATP and releases CO₂ without making any sugar. This is photorespiration.
Photorespiration is worst on hot, dry, bright days. To save water, plants close their stomata; CO₂ inside the leaf falls and O₂ from the light reactions builds up, so rubisco grabs O₂ more often. In many crop plants, photorespiration can waste a significant share of the carbon fixed.
C₄ and CAM plants: two solutions
C₄ plants (corn, sugar cane, and many warm-season grasses such as big bluestem of the tallgrass prairie) separate the steps in space:
- In mesophyll cells, the enzyme PEP carboxylase, which does not bind O₂, fixes CO₂ into a four-carbon acid.
- The acid moves into bundle-sheath cells around the veins and releases CO₂ there, keeping CO₂ high around rubisco.
- The Calvin cycle runs in the bundle-sheath cells with little photorespiration.
This costs extra ATP, but in hot, sunny conditions C₄ plants can be more productive and use water more efficiently. That's one reason grain corn grows so well in the warm summers of southwestern Ontario.
CAM plants (pineapple, cacti, jade plants and many succulents) separate the steps in time:
- At night, they open their stomata (when it is cool and less water is lost) and fix CO₂ into four-carbon acids stored in the vacuole.
- During the day, stomata stay closed to save water; the stored acids release CO₂ for the Calvin cycle.
Most plants, including wheat, soybeans, potatoes and maple trees, are C₃ plants: the first stable product of CO₂ fixation is the three-carbon 3-PGA.
Worked exampleLet's do Calvin cycle accounting
A leaf fixes 18 molecules of CO₂ in the Calvin cycle. How many turns of the cycle are needed? How many RuBP, ATP and NADPH are used? How many net G3P and glucose molecules can be made?
- One turn fixes one CO₂, so 18 turns.
- RuBP used: one per CO₂ = 18 RuBP (all regenerated).
- Per 3 CO₂: 9 ATP and 6 NADPH. 18 CO₂ = 6 groups of 3: ATP = 6 × 9 = 54; NADPH = 6 × 6 = 36.
- Net G3P: 18 ÷ 3 = 6 G3P; glucose: 6 ÷ 2 = 3 glucose.
- Check carbon: 18 C fixed = 6 G3P × 3 C = 18 C ✔
18 turns, 18 RuBP (regenerated), 54 ATP, 36 NADPH → 6 G3P → 3 glucose.
Worked exampleLet's interpret a ¹⁴C experiment
Algae are given ¹⁴CO₂ in the light and then the light is suddenly switched off, while CO₂ supply continues. Predict what happens to the levels of 3-PGA and RuBP over the next minute, and explain.
- In the dark, the light reactions stop, so ATP and NADPH run out quickly.
- Without them, 3-PGA can't be reduced to G3P: 3-PGA builds up.
- Without G3P (and ATP), RuBP can't be regenerated, but rubisco keeps using the RuBP that is there to fix CO₂: RuBP falls.
- This is the result Calvin's group observed, and it helped confirm that RuBP is the CO₂ acceptor and 3-PGA its product.
3-PGA rises and RuBP falls after the light is switched off, showing that the reduction and regeneration steps depend on ATP and NADPH from the light reactions.
Just for funRubisco is the world's most abundant protein, but also one of the slowest enzymes at its job and easily confused by oxygen. If it were an employee, it would be the one who shows up everywhere, works at a snail's pace, and sometimes files the wrong paperwork. Life on Earth depends on it anyway.
Hands-on activityStarch-free leaves: does sugar come from photosynthesis?
You need: a healthy potted geranium or other soft-leaved houseplant, aluminum foil, a paper clip, and either the iodine test at school (see the next lesson) or the observation alternative below.
Safety note: the starch test needs boiling alcohol and iodine solution, so it is done only at school with a teacher. At home, do the set-up and a simple observation version.
- Put the plant in a dark cupboard for 48 hours. (The leaves use up their stored starch.)
- Fold a strip of foil over part of one leaf (top and bottom), held with a paper clip, so that part of the leaf gets no light. Put the plant in a bright window for 1 to 2 days.
- At school: the teacher removes the leaf, takes off the foil, removes the chlorophyll with hot alcohol in a water bath, and adds iodine: starch turns blue-black.
- At home alternative: after a week with the foil on, compare the covered and uncovered parts of the leaf. The covered part may become paler.
- Draw and label the leaf before and after. Write a prediction for the iodine test result.
What to look for: only the parts of the leaf that received light make starch: the uncovered part turns blue-black with iodine, the covered strip stays brown-yellow. This shows that the sugar stored as starch is made by photosynthesis, which needs light (for ATP and NADPH used in the Calvin cycle). If the covered patch turns paler at home, that shows leaves need light to keep making chlorophyll.
Your turn
1Name the three phases of the Calvin cycle.
Carbon fixation, reduction, and regeneration of RuBP.
It fixes CO₂ by attaching it to RuBP, producing an unstable 6C molecule that splits into two 3-PGA.
3How many ATP and NADPH are needed to make one G3P?
9 ATP and 6 NADPH (from 3 CO₂).
4How many CO₂, ATP and NADPH are needed for one glucose?
5Why is only one of every six G3P made "profit" for the plant?
Five of the six G3P (15 C) must be used to regenerate three RuBP (15 C) so the cycle can continue; only one G3P (3 C, equal to the 3 CO₂ fixed) is a net gain.
6What is photorespiration and when is it most common?
Rubisco binds O₂ instead of CO₂, wasting ATP and releasing CO₂ without making sugar. It's most common on hot, dry, bright days when stomata close and CO₂ inside the leaf is low.
7How do C₄ plants reduce photorespiration?
They fix CO₂ with PEP carboxylase (which doesn't bind O₂) into a 4C acid in mesophyll cells, then release CO₂ in bundle-sheath cells, keeping CO₂ high around rubisco.
More practice: warm-up
1How many carbons are in RuBP?
2What is the first stable product of CO₂ fixation in C₃ plants?
3-PGA (3-phosphoglycerate).
3Where in the chloroplast is the Calvin cycle?
4Give one example of a C₄ plant and one of a CAM plant.
C₄: corn (or sugar cane). CAM: pineapple (or a cactus, jade plant).
5Who led the team that worked out the Calvin cycle?
Melvin Calvin (with Andrew Benson and James Bassham).
More practice: on your own
1Explain why the Calvin cycle stops soon after a plant is put in the dark, even though it doesn't use light directly.
It depends on a continuous supply of ATP and NADPH from the light reactions, which stop in the dark. Some Calvin cycle enzymes are also activated by light. Without ATP and NADPH, 3-PGA can't be reduced and RuBP can't be regenerated.
2Why do CAM plants open their stomata at night instead of during the day?
Nights are cooler and more humid, so much less water is lost through open stomata. The CO₂ is stored as acids and used during the day while stomata stay closed, which suits hot, dry habitats.
3A plant's leaf releases more CO₂ than expected on a hot, sunny afternoon, even though it is photosynthesizing. Explain.
Stomata are partly closed to save water, so CO₂ in the leaf falls and O₂ rises. Rubisco binds O₂ more often: photorespiration, which releases CO₂. (The plant's own cellular respiration also releases CO₂.)
4Trace the path of a carbon atom from CO₂ in the air to sucrose in a maple's phloem.
CO₂ enters through stomata → diffuses to a mesophyll chloroplast → fixed by rubisco to RuBP → 3-PGA → reduced to G3P (using ATP, NADPH) → G3P exported → combined to glucose and fructose → joined into sucrose → loaded into the phloem.
5Explain why it makes sense that C₄ photosynthesis costs extra ATP but still benefits corn in a hot Ontario summer.
Pumping CO₂ into bundle-sheath cells costs extra ATP, but in hot, sunny conditions photorespiration in C₃ plants wastes even more energy and carbon. C₄ plants avoid most photorespiration and can keep stomata less open (saving water) while still fixing CO₂ quickly. In bright light, ATP is plentiful, so the extra cost is affordable.
Challenge
1Show that the Calvin cycle accounts for all carbon atoms when 6 CO₂ are fixed.
6 CO₂ + 6 RuBP (6 × 5 = 30 C) → 12 3-PGA (36 C) → 12 G3P (36 C). Of these, 10 G3P (30 C) regenerate 6 RuBP (30 C), and 2 G3P (6 C) are the net product = 1 glucose (6 C). Carbon in = 6 (from CO₂) + 30 (from RuBP) = 36 = carbon out (30 in RuBP + 6 in product). ✔
2If each ATP holds 30.5 kJ/mol and each NADPH about 220 kJ/mol, estimate the energy input to make one mole of glucose in the Calvin cycle and the efficiency of storing it (glucose = 2870 kJ/mol).
Energy in: 18 × 30.5 + 12 × 220 = 549 + 2640 = 3189 kJ. Efficiency = 2870 ÷ 3189 × 100% ≈ 90%. The Calvin cycle itself stores energy efficiently; most of the losses in photosynthesis happen earlier, when light energy is converted (much becomes heat) and in photorespiration.
3Rising atmospheric CO₂ is expected to affect C₃ and C₄ plants differently. Predict which type would benefit more from higher CO₂ levels, and explain using photorespiration.
C₃ plants would generally benefit more. Higher CO₂ increases the ratio of CO₂ to O₂ at rubisco, reducing photorespiration and speeding up carbon fixation. C₄ plants already concentrate CO₂ around rubisco, so extra atmospheric CO₂ helps them less. (Other factors such as temperature, water and nutrients also matter.)
Quiz
1. Which enzyme fixes CO₂ in the Calvin cycle?
2. How many ATP and NADPH does the Calvin cycle use to make one molecule of glucose?
3. In the Calvin cycle, NADPH is used during:
4. Photorespiration occurs when:
5. CAM plants reduce water loss by:
6. In Calvin's experiments, which compound became radioactive first after a very short exposure to ¹⁴CO₂?
Big idea: The Calvin cycle (C3.2, C2.1) uses ATP and NADPH from the light reactions to fix and reduce CO₂ into G3P. Students should be able to do the carbon and energy accounting (9 ATP and 6 NADPH per G3P) and explain photorespiration and the C₄/CAM adaptations (C1.1 links to environments and crops).
How to help: Ask your teen to explain the cycle using carbon counts: 3 CO₂ + 3 RuBP (15 C) → 6 3-PGA (18 C) → 6 G3P → 1 out, 5 recycled. The leaf-covering activity needs a few days; the iodine starch test is done only at school because it uses boiling alcohol.
Common difficulties:
- Thinking the Calvin cycle runs at night ("dark reactions").
- Thinking each turn makes a glucose; it takes 6 turns (6 CO₂) per glucose.
- Mixing up C₄ (separate cells: space) and CAM (night/day: time).
- Forgetting that plants also do cellular respiration.