The effect of plastid pigments and respiration intensity on crop yield

The efficiency of the leaf apparatus and the root system directly determines future yield. It is important for an agronomist to understand how these processes limit crop development in the field. Correct regulation of sowing density and nitrogen nutrition helps manage the balance of photosynthesis and respiration.

• Chlorophyll content — 2–6 mg per 0.01 m² of leaf area
• Share of chlorophyll in leaf dry matter — 0.5–2.0 %

Pigments and Photosynthetic Intensity

The accumulation of plastid pigments (chlorophylls a and b, carotenoids) in chloroplasts creates the basis for photosynthesis. However, there is no direct proportional relationship between the volume of chlorophyll and the rate of photosynthesis. The efficiency of pigment function is assessed through the assimilation number — the quantity of milligrams of CO₂ assimilated per unit of chlorophyll. As the chlorophyll content increases from 0.7 to 15.6 mg/g of fresh mass (more than 20-fold), the intensity of photosynthesis increases only 2-fold, and the assimilation number drops 10-fold.

Chlorophyll deficiency begins to strictly limit photosynthesis only in two unfavorable situations. This occurs during nitrogen deficiency in the soil or in overly dense crops where plants experience a shortage of sunlight. The rate of dry matter accumulation also depends heavily on the type of photosynthesis of a particular crop.

Respiration as a Driver of Mineral Nutrition

Plant respiration is the oxidative breakdown of organic substances with the consumption of oxygen and the release of carbon dioxide. The rate of this process is called respiration intensity. It is determined by the volume of oxygen absorbed or carbon dioxide released per unit of time per 1 g of dry matter. Respiration provides the plant with energy and directly triggers the process of root nutrition.

During respiration, hydrogen cations and carbonic acid anions are formed. They serve as an exchange pool, thanks to which roots absorb nutrients from the soil. The released energy is used for the active transport of ions through the cell membrane (plasmalemma). Furthermore, root exudates during respiration contain weak organic acids that convert poorly accessible mineral soil elements into a mobile, easily digestible form for plants.

The oxidative breakdown of glucose during respiration is expressed by the following chemical equation:

С₆Н₁₂О₆ + 6О₂ → 6СО₂ + 6Н₂О + 2875 kJ

All respiration reactions are closely linked to processes of synthesis and the construction of cellular structures. Intermediate compounds arising from the breakdown of sugars are used by cells as building material. By regulating growing conditions through agricultural practices, it is possible to create an optimal respiration balance and direct the energy of respiration toward forming a high yield.

Respiration is the main energy engine of a plant, providing energy for all processes from cell growth to the uptake of nutrients from the soil. During the complete oxidation of one gram-molecule of hexose, 6 molecules of oxygen are consumed, an equivalent amount of carbon dioxide and water is released, and 2875 kJ of energy is liberated. This energy is not released instantaneously but is stored in stages in the form of ATP molecules. The entire respiration process is divided into three consecutive stages: glycolysis, the Krebs cycle, and the electron transport chain (ETC).

• Oxidation energy of 1 g-mol of hexose — 2875 kJ
• Oxygen consumption per 1 g-mol — 6 molecules
• Energy yield from fermentation — 2 ATP molecules

Glycolysis: Nine Steps of Anaerobic Breakdown

Glycolysis, or the Embden-Meyerhof-Parnas pathway, is localized in the cell cytoplasm and occurs without the participation of oxygen. During this process, a glucose molecule is broken down into pyruvic acid (pyruvate). The entire pathway consists of nine strictly sequential enzymatic reactions, each of which prepares the substrate for energy extraction.

1. Glucose phosphorylation. Hexokinase transfers a phosphate group from an ATP molecule to glucose, forming glucose-6-phosphate (the reaction requires ATP energy expenditure, converting it into ADP).
2. Isomerization. Phosphoglucoisomerase converts the six-carbon ring of glucose-6-phosphate into the five-carbon ring of fructose-6-phosphate.
3. Second phosphorylation. Phosphofructokinase attaches another phosphate group from ATP to the first carbon atom of fructose-6-phosphate, forming fructose-1,6-diphosphate.
4. Cleavage into trioses. Aldolase splits fructose-1,6-diphosphate into dihydroxyacetone phosphate and glyceraldehyde-3-phosphate. At the same time, dihydroxyacetone phosphate is completely converted into glyceraldehyde-3-phosphate under the action of phosphotriose isomerase.
5. Oxidation with NAD reduction. Two molecules of glyceraldehyde-3-phosphate are oxidized under the action of dehydrogenase, giving up hydrogen to reduce NAD to NADH2, and are converted into two molecules of glycerate-1,3-bisphosphate (the first energy-storage reaction).
6. First ATP synthesis. Glycerate-3-phosphate kinase transfers a phosphate group from glycerate-1,3-bisphosphate to ADP, forming ATP and glycerate-3-phosphate (this reaction stimulates the progression of previous stages).
7. Intramolecular transfer. Phosphoglycerate mutase converts glycerate-3-phosphate into glycerate-2-phosphate.
8. Dehydration. Enolase splits off a water molecule from glycerate-2-phosphate, converting it into phosphoenolpyruvic acid (phosphoenolpyruvate) with the formation of a high-energy phosphate bond.
9. Pyruvate formation. Pyruvate kinase transfers a phosphate group from phosphoenolpyruvate to ADP, forming ATP and unstable enolpyruvic acid, which converts into stable pyruvate (pyruvic acid).

Respiratory fork: aerobic oxidation or fermentation

The formation of pyruvic acid completes glycolysis, but the further fate of pyruvate depends on the presence of oxygen in plant tissues. With good soil aeration, pyruvic acid freely diffuses into the mitochondria. These organelles, with their double membrane and internal folds (cristae), contain a complete set of enzymes to complete the respiratory cycle.

In the mitochondria, the di- and tricarboxylic acid cycle, known as the Krebs cycle, is triggered. Here, pyruvic acid is completely oxidized to carbon dioxide and water, completing the breakdown of carbohydrates, fats, and proteins. This pathway is most beneficial for the plant, as it provides the maximum amount of energy in the form of ATP.

If oxygen access is restricted — for example, due to field flooding, soil compaction, or crust formation — respiration shifts to an emergency mode. Pyruvate cannot be oxidized in the mitochondria and remains in the cytoplasm. The plant is forced to use a backup anaerobic pathway to obtain energy.

The oxidation of pyruvic acid occurs in two stages: 1) oxidative decarboxylation of pyruvate to acetyl-CoA and 2) oxidation of the acetyl-CoA residue in the Krebs cycle. Oxidative decarboxylation of pyruvic acid is carried out with the participation of the pyruvate dehydrogenase multienzyme complex, which consists of three enzymes and five coenzymes. The coenzymes are thiamine pyrophosphate (TPP) – a phosphorylated derivative of vitamin B1, lipoic acid, coenzyme A, FAD, and NAD. Pyruvic acid interacts with the enzyme thiamine pyrophosphate decarboxylase. In this process, CO2 is split off and a hydroxyethyl derivative of TPP is formed, which reacts with the oxidized form of lipoic acid. The disulfide bond of lipoic acid is broken and an oxidation-reduction reaction occurs: the hydroxyethyl group attached to one sulfur atom is oxidized to an acetyl group, and the second sulfur atom of lipoic acid is reduced. The resulting acetyllipoic acid interacts with coenzyme A, creating acetyl-CoA and the reduced form of lipoic acid.

As a result of the oxidative decarboxylation of pyruvic acid, acetyl-CoA, CO2, and NADH are formed. Acetyl-CoA is involved directly in the Krebs cycle, which occurs during its interaction with oxaloacetic acid. In this reaction, under the influence of the enzyme citrate synthase, citric acid is formed and a CoA molecule is released. The process of citric acid synthesis becomes possible due to the energy of pyruvate oxidation. Subsequent transformations in the cycle involve not citric acid, but isocitric acid, which is formed from citric acid via the intermediate formation of cis-aconitic acid.

The reactions of converting citric acid into cis-aconitic acid and further into isocitric acid are catalyzed by aconitase. Isocitric acid, under the influence of NAD-dependent isocitrate dehydrogenase, is oxidized into an unstable compound – oxalosuccinic acid, which is immediately decarboxylated to form α-ketoglutaric acid. During this process, NADH is formed, as well as the first CO2 molecule in the cycle and the second since the start of pyruvic acid metabolism. Next, during the oxidative decarboxylation of α-ketoglutaric acid

Discovered in 1937 by English biochemists H. Krebs and W. Johnson.

With the help of the α-ketoglutarate dehydrogenase complex, NADH is formed, succinyl-CoA in the presence of mineral phosphorus (Pi) and ADP is converted into succinic acid (succinate), which is accompanied by the formation of ATP in the process of substrate-level phosphorylation. Next, succinic acid, under the influence of succinate dehydrogenase, the coenzyme of which is FAD, is converted into fumaric acid with the formation of FADH2. During the reaction, fumaric acid, reacting with water in the presence of the enzyme fumarate hydratase, forms malic acid (malate), which is then oxidized to oxaloacetic acid with the help of NAD-dependent malate dehydrogenase. The latter spontaneously converts into the enol form, reacts with the next acetyl-CoA molecule, and the cycle repeats again. The first stage of pyruvic acid cleavage and the reactions of the Krebs cycle occur in the mitochondrial matrix.

Fig. 39. The Krebs cycle (tricarboxylic acid cycle)

Thus, the following transformations occur during the Krebs cycle:

• Consumed: one molecule each of pyruvate, ADP, and FAD, four NAD, and two water molecules.
• Synthesized: four NADH molecules, one ATP, one FADH2, and two water molecules.
• Released: three CO2 molecules.

The Krebs cycle functions only under aerobic* conditions, although oxygen does not directly participate in its reactions. Glycolysis reactions also proceed without oxygen, yet it is known that oxygen consumption is a characteristic feature of the respiration process. The need for oxygen arises at the final stage of the respiration process, associated with the oxidation of reduced nicotinamide adenine dinucleotide (NADH) and flavin adenine dinucleotide (FADH2) coenzymes, which contain a significant portion of the energy previously held in the hexose in the form of electrons.

*Aerobic – requiring the presence of molecular oxygen.

Electron transport chain. Energy release occurs through the transfer of electrons from NADH and FADH2 along the electron transport chain (ETC) to free oxygen, which serves as the terminal electron acceptor. The first electron carrier in the ETC is NAD; the second is flavin enzymes; the third is cytochromes. At the end of the chain, electrons are captured by oxygen and combined with protons (hydrogen ions) to form a water molecule.

According to current concepts, the mitochondrial respiratory ETC includes four main multi-enzyme complexes and two components of low molecular weight:

• Ubiquinone – a lipid consisting of benzoquinone connected to a long hydrophobic isoprenoid side chain.
• Cytochrome C – a protein molecule containing a heme-iron-porphyrin ring (Fig. 40; Sheudzhen A.Kh., 2000).

Fig. 40. Mitochondrial respiratory electron transport chain

How respiration provides the plant with energy: ATP balance

Plant respiration is a multi-step process of converting carbohydrates into a form accessible to cells. Everything begins with the breakdown of glucose into two molecules of pyruvate (pyruvic acid). Then, in the mitochondria, pyruvate is converted into acetyl groups, which are involved in the Krebs cycle. Here, they are sequentially oxidized to carbon dioxide, and the released electrons are transferred via the electron transport chain (ETC) to oxygen, forming water.

The electron transport chain is localized in the inner mitochondrial membrane and operates like a multi-stage conveyor. Electron transfer and the associated transport of hydrogen ions are carried out by four large protein complexes:

• Complex I (NADH:ubiquinone oxidoreductase) carries out the transfer of electrons from intramitochondrial NADH to ubiquinone Q. The complex is FMN-dependent and contains three iron-sulfur centers (FeSN1-3).
• Complex II (succinate:ubiquinone oxidoreductase) catalyzes the oxidation of succinate by ubiquinone. This reaction is FAD-dependent and proceeds with the participation of three iron-sulfur centers (FeSS1-3).
• Complex III (ubiquinone:cytochrome C oxidoreductase) transfers electrons from reduced ubiquinone to cytochrome C. Its composition includes cytochromes b556, b560, c1 and an iron-sulfur protein.
• Complex IV (cytochrome oxidase) — the terminal section of the chain, where electrons are transferred from cytochrome a-CuA to cytochrome a3-CuB, and then to oxygen. It contains four redox components: cytochromes a and a3, as well as two copper atoms.

During the complete aerobic oxidation of one glucose molecule, the plant receives 38 ATP molecules. Almost all of them, except for two, are formed during mitochondrial reactions. In this process, the majority of the energy (all ATP, except for four molecules) is synthesized during the oxidation of NADH2 and FADH2. A detailed energy balance by stage is presented in the table.

Phosphorylation mechanism and real respiration efficiency

To explain the mechanism of oxidative phosphorylation, three theories have been proposed: chemical, conformational, and chemiosmotic. The most widely accepted today is the chemiosmotic theory. Its basis lies in the concept that ATP synthesis occurs due to the difference in proton concentrations on both sides of the inner mitochondrial membrane.

According to this concept, during electron transport along the membrane, protons (H+ ions) are pumped out of the mitochondrial matrix into the external intermembrane space. They can return to the matrix only through a special channel containing the enzyme complex ATP-synthetase (ATPase). The resulting electrochemical gradient is the driving force that compels the enzyme to synthesize ATP from ADP and inorganic phosphate.

• ATP bond (third phosphate ester) — 42 kJ/mol (10 kcal/mol)
• Energy of 38 ATP molecules — 1591 kJ/mol (380 kcal/mol)
• Free energy of glucose oxidation — 2872 kJ/mol (686 kcal/mol)
• Theoretical efficiency of energy storage — 55 %