outline the three main stages of the Calvin cycle: rubisco catalyses the fixation of carbon dioxide by combination with a molecule of ribulose bisphosphate (RuBP), a 5C compound, to yield two molecules of glycerate 3-phosphate (GP), a 3C compound, GP

Photosynthesis – Energy Transfer Process (Cambridge 9700)

Learning Objectives

• Describe the three main stages of the Calvin (light‑independent) cycle and the role of Rubisco.
• Explain how the light‑dependent reactions provide the ATP and NADPH required for the Calvin cycle, including the proton‑gradient mechanism of photophosphorylation.
• Identify the major photosynthetic pigments, their absorption maxima and how action spectra are obtained.
• Analyse the stoichiometric relationships (ATP/NADPH per CO₂) and the link to the respiratory quotient (RQ≈1).
• Discuss photorespiration and the main environmental factors that limit photosynthesis.
• Outline practical investigations that test these limiting factors and relate the topic to other Cambridge syllabus areas.

1. Chloroplast Ultrastructure – Linking Structure to Function

The chloroplast is divided into two functional compartments:

• Thylakoid system (granum & lamellae) – site of the light‑dependent reactions.

  • Thylakoid membranes contain Photosystem II, the cytochrome b₆f complex, Photosystem I and ATP synthase.
  • Stacks of thylakoids (grana) increase surface area for light capture.

• Stroma – aqueous matrix surrounding the thylakoids where the Calvin cycle occurs.

  • Contains enzymes of carbon fixation, the enzymes of the reduction and regeneration phases, and soluble ribulose‑1,5‑bisphosphate (RuBP).

2. Light‑Dependent Reactions (Thylakoid Phase)

2.1 Overview

Water is split, electrons are excited, and a proton gradient (ΔpH) is generated across the thylakoid membrane. The gradient drives ATP synthase (photophosphorylation) while NADP⁺ is reduced to NADPH.

2.2 Step‑by‑step

2.3 Energy Yield per 3 CO₂ Fixed

To sustain one full turn of the Calvin cycle (3 CO₂), the light‑dependent reactions must supply:

• 9 ATP (3 ATP per CO₂)
• 6 NADPH (2 NADPH per CO₂)

These values are derived from the stoichiometry of the three Calvin‑cycle stages (see Section 4).

3. Photosynthetic Pigments & Action Spectra

Action spectrum: By measuring the rate of O₂ evolution (or DCPIP reduction) at different wavelengths, students can plot an action spectrum that mirrors the combined absorption spectra of these pigments, confirming which wavelengths drive photosynthesis most efficiently.

4. Calvin Cycle – Light‑Independent Reactions (Stroma)

4.1 Overall View

Three CO₂ molecules are fixed, consuming 9 ATP and 6 NADPH, to produce one net glyceraldehyde‑3‑phosphate (G3P). Six G3P molecules are generated; five are recycled to regenerate RuBP, and one can leave the cycle for biosynthesis (e.g., glucose formation).

4.2 Stage 1 – Carbon Fixation

Enzyme: Ribulose‑1,5‑bisphosphate carboxylase/oxygenase (Rubisco)

\$\text{RuBP (5C)} + \text{CO}_2 \xrightarrow{\text{Rubisco}} 2\,\text{3‑PGA (GP, 3C)}\$

• No ATP or NADPH required.
• Each CO₂ yields two molecules of 3‑phosphoglycerate (3‑PGA, also called GP).

4.3 Stage 2 – Reduction

Overall for one CO₂:

\$\text{3‑PGA} + \text{ATP} + \text{NADPH} \longrightarrow \text{G3P} + \text{ADP} + P_i + \text{NADP}^+\$

4.4 Stage 3 – Regeneration of RuBP

Five of the six G3P molecules are rearranged through a series of aldol condensations, isomerisations and dephosphorylations to regenerate three RuBP molecules.

4.5 Stoichiometry Recap

4.6 Worked Example – Scaling the Cycle

If a leaf fixes 6 CO₂ (i.e., two full turns of the cycle):

• ATP required = 9 ATP × 2 = 18 ATP
• NADPH required = 6 NADPH × 2 = 12 NADPH
• Net G3P produced = 1 G3P × 2 = 2 G3P (which can be used to form one glucose molecule after further reactions).

5. Photorespiration – An Alternative Rubisco Reaction

Rubisco can also add O₂ to RuBP, producing one molecule of 3‑PGA and one molecule of 2‑phosphoglycolate (a toxic 2‑C compound). The latter must be recycled via the photorespiratory pathway, consuming ATP and releasing CO₂, thus lowering photosynthetic efficiency.

• Occurs when O₂ levels are high and CO₂ is low (e.g., hot, dry conditions when stomata close).
• Reduces the net gain of carbohydrate per CO₂ fixed.

6. Environmental Factors that Limit Photosynthesis

7. Practical Investigations (AO3)

• Leaf‑disc floating assay – Vary light intensity, wavelength (using coloured filters), CO₂ concentration, or temperature; measure disc rise time as a proxy for O₂ evolution.
• DCPIP reduction assay – Extract chlorophyll, add DCPIP, and record the rate of colour loss under different light qualities to construct an action spectrum.
• Gas‑evolution apparatus – Use a sealed chamber with a gas‑sensor to measure O₂ production and CO₂ uptake while systematically altering one factor (e.g., temperature 10‑40 °C) to determine limiting ranges.
• Photorespiration test – Compare O₂ evolution under normal air (≈21 % O₂) with that under low O₂ (e.g., N₂‑flushed) to demonstrate the impact of oxygenation.

8. Cross‑Topic Links (Cambridge Syllabus)

• Cellular Respiration (Topic 12) – NADPH from the light reactions is analogous to NADH from glycolysis; both provide reducing power for biosynthesis. The photosynthetic RQ≈1 (CO₂ fixed = O₂ released) mirrors the respiratory RQ for carbohydrate oxidation.
• Transport in Plants (Topic 9) – Water required for photolysis is supplied via the xylem; O₂ and CO₂ exchange occurs through stomata, linking gas exchange to transpiration.
• Energy & Metabolism (Topic 13) – The Calvin cycle stores solar energy as chemical energy (carbohydrates), which is later released during cellular respiration.

9. Summary Table – Calvin Cycle at a Glance

10. Suggested Diagram

A circular diagram should show:

• RuBP → 3‑PGA → G3P → RuBP (with arrows indicating the three stages).
• Enzyme names placed on each arrow.
• Inputs of ATP (green arrows) and NADPH (blue arrows) entering the reduction and regeneration phases.
• An inset of the thylakoid stack illustrating PS II → ETC → PS I → ATP synthase, with the proton gradient highlighted.