11.7 Where are the ATP and NADPH Used?

We learnt that the products of light reaction are ATP, NADPH and \(O _2\). Of these \(O _2\) diffuses out of the chloroplast while ATP and NADPH are used to drive the processes leading to the synthesis of food, more accurately, sugars. This is the biosynthetic phase of photosynthesis. This process does not directly depend on the presence of light but is dependent on the products of the light reaction, i.e., ATP and NADPH, besides \(CO _2\) and \(H _2 O\). You may wonder how this could be verified; it is simple: immediately after light becomes unavailable, the biosynthetic process continues for some time, and then stops. If then, light is made available, the synthesis starts again.

Can we, hence, say that calling the biosynthetic phase as the dark reaction is a misnomer? Discuss this amongst yourselves.

Let us now see how the ATP and NADPH are used in the biosynthetic phase. We saw earlier that \(CO _2\) is combined with \(H _2 O\) to produce \(\left( CH _2 O \right)_n\) or sugars. It was of interest to scientists to find out how this reaction proceeded, or rather what was the first product formed when \(CO _2\) is taken into a reaction or fixed. Just after world war II, among the several efforts to put radioisotopes to beneficial use, the work of Melvin Calvin is exemplary. The use of radioactive \({ }^{14} C\) by him in algal photosynthesis studies led to the discovery that the first \(CO _2\) fixation product was a 3 -carbon organic acid. He also contributed to working out the complete biosynthetic pathway; hence it was called Calvin cycle after him. The first product identified was 3-phosphoglyceric acid or in short PGA. How many carbon atoms does it have?

Scientists also tried to know whether all plants have PGA as the first product of \(CO _2\) fixation, or whether any other product was formed in other plants. Experiments conducted over a wide range of plants led to the discovery of another group of plants, where the first stable product of \(CO _2\) fixation was again an organic acid, but one which had 4 carbon atoms in it. This acid was identified to be oxaloacetic acid or OAA. Since then \(CO _2\) assimilation during photosynthesis was said to be of two main types: those plants in which the first product of \(CO _2\) fixation is a \(C _3\) acid (PGA), i.e., the \(C _3\) pathway, and those in which the first product was a \(C _4\) acid (OAA), i.e., the \(C _4\) pathway. These two groups of plants showed other associated characteristics that we will discuss later.

The Primary Acceptor of \(\mathrm{CO}_2\)

Let us now ask ourselves a question that was asked by the scientists who were struggling to understand the ‘dark reaction’. Howmany carbon atoms would a molecule have which after accepting (fixing) \(CO _2\), would have 3 carbons (of PGA)?

The studies very unexpectedly showed that the acceptor molecule was a 5-carbon ketose sugar – ribulose bisphosphate (RuBP). Did any of you think of this possibility? Do not worry; the scientists also took a long time and conducted many experiments to reach this conclusion. They also believed that since the first product was a \(C _3\) acid, the primary acceptor would be a 2-carbon compound; they spent many years trying to identify a 2-carbon compound before they discovered the 5-carbon RuBP.

The Calvin Cycle

Calvin and his co-workers then worked out the whole pathway and showed that the pathway operated in a cyclic manner, the RuBP was regenerated. Let us now see how the Calvin pathway operates and where the sugar is synthesised. Let us at the outset understand very clearly that the Calvin pathway occurs in all photosynthetic plants; it does not matter whether they have \(C _3\) or \(C _4\) (or any other) pathways (Figure 11.8).

For ease of understanding, the Calvin cycle can be described under three stages: carboxylation, reduction and regeneration.

• Carboxylation-Carboxylation is the fixation of \(CO _2\) into a stable organic intermediate. Carboxylation is the most crucial step of the Calvin cycle where \(CO _2\) is utilised for the carboxylation of RuBP . This reaction is catalysed by the enzyme RuBP carboxylase which results in the formation of two molecules of 3-PGA. Since this enzyme also has an oxygenation activity it would be more correct to call it RuBP carboxylase-oxygenase or RuBisCO.
• Reduction – These are a series of reactions that lead to the formation of glucose. The steps involve utilisation of 2 molecules of ATP for phosphorylation and two of NADPH for reduction per \(CO _2\) molecule fixed. The fixation of six molecules of \(CO _2\) and 6 turns of the cycle are required for the formation of one molecule of glucose from the pathway.
• Regeneration – Regeneration of the \(CO _2\) acceptor molecule RuBP is crucial if the cycle is to continue uninterrupted. The regeneration steps require one ATP for phosphorylation to form RuBP.

Hence for every \(CO _2\) molecule entering the Calvin cycle, 3 molecules of ATP and 2 of NADPH are required. It is probably to meet this difference in number of ATP and NADPH used in the dark reaction that the cyclic phosphorylation takes place.

To make one molecule of glucose 6 turns of the cycle are required. Work out how many ATP and NADPH molecules will be required to make one molecule of glucose through the Calvin pathway.

It might help you to understand all of this if we look at what goes in and what comes out of the Calvin cycle.
\(
\begin{array}{|c|c|}
\hline \text { In } & \text { Out } \\
\hline Six ~CO_2 & \text { One glucose } \\
\hline 18 \text { ATP } & 18 \text { ADP } \\
\hline 12 \text { NADPH } & 12 \text { NADP } \\
\hline
\end{array}
\)