Synthesis of ATP Formation:

In the early 1930s it was clear that the ADP phosphorylation was coupled to respiration. Lehninger and other workers between 1948-51 conducted biochemical studies on isolated mitochondria and tissue samples.

From their studies following general inferences emerged:

i. Phosphorylation depended upon oxygen.

ii. Mitochondria when isolated catalyzed oxygen-dependent phosphorylation in collaboration with oxidations of intermediates of Krebs cycle.

iii. When NADH+ was added to the broken mitochondria, it was immediately reduced to NAD+.

ATP molecule is synthesized in a cell during various exergonic reactions, e.g., during respiratory metabolism and electron transport in the chloroplast during photosynthesis, which consists of sequential and regulated chemical steps. ATP consists of three molecular species, nucleoside adenosine, D-ribose sugar and three phosphates attached to the D-ribose sugar (Fig. 17-10).

Three phosphates are connected via acid anhydride linkages and are termed “high energy” bonds which are represented by the symbol P. When the terminal phosphate is cleaved the energy released is approximately 7.3 kcal/mole under physiological conditions.

The “high energy” nature of this molecule is attributed to resonance stabilization of the hydrolysis products and intra-molecular electrostatic repulsion. ATP is a versatile carrier of chemical energy and is cleaved in four different ways and the point at which ATP is cleaved is determined by specific enzymes.

The physiological mechanisms involved in the synthesis of ATP have now thoroughly been investigated. The chloroplasts and mitochondria are the two organelles in the cell which synthesize ATP. Most of the ATP synthesized by the chloroplasts of green plant cells comes from the photophosphorylation involving the photolysis of water with the release of hydrogen atoms or electrons and protons.

The electrons pass through different electrons carrier molecules arranged in a down gradient sequence as far as their reduction oxidation potential is concerned so as to make efficient energy transfer in cellular metabolism.

In the ultimate reaction electrons and protons help in the reduction of carbon dioxide to form organic molecules. Most of the ATP synthesized by the chloroplasts are utilized in efficient running of the process of photosynthesis.

Mitochondria are supposed to be the biggest power houses with immense capacity to synthesize ATP (Fig. 17-10a). The energy in the organic molecules after oxidation passes the electrons through the carrier molecules and get ultimately accepted by oxygen forming water.

It is very puzzling how the electron transfer through series of carrier molecules is coupled with the synthesis of ATP. ATP formation process is coupled to transport of electrons along the cytochrome chain towards molecular oxygen and is said to be oxidative phosphorylation.

The normal site of oxidative phosphorylation in plant and animal cells is mitochondria. ATP production also requires oxygen in the last step of the series and is, thus, referred to as oxidative phosphorylation. In comparison during photosynthesis there also occurs phosphorylation of ADP resulting in ATP formation. This is called photophosphorylation and is light dependent.

ATP yield from oxidative phosphorylation is usually expressed as P/O ratio, i.e., moles of inorganic phosphate recovered in organic form per oxygen atom consumed. On an average 2-3 ATP molecules are formed for a particular substrate (e.g., organic acid).

Thus while during pyruvate to acetyl-CoA the P/O ratio is 3, during α-ketoglutarate to succinate it is 4. Specific chemical uncoupling agents like 2, 3-dinitrophenol has been used to dissociate ADP oxidative phosphorylation

from respiration. Thus ATP synthesis is inhibited but the rate of electron transport is not affected. Alternatively energy transfer inhibitors e.g., oligomycin, is used. This inhibits oxidative phosphorylation and also the electron transport rate.

Hinkle and McCarty (1978) have described in detail how cells make ATP. For full details a reference may be made to their review.

With the available information, it is now possible to calculate the molecules of ATP formed during different steps.

A brief account is given below:

In summary, it may be mentioned that during aerobic glycolysis, 8 ATP molecules are produced while during pyruvic acid oxidation and Krebs cycle, a total of 24 ATP molecules are yielded.

In the following summary equations of the different stages of cell respiration are given:

Glycolysis:

Glucose + 2ADP + 2Pi → 2 pyruvic acid + 2 ATP + 4 H

Pyruvic acid oxidation:

2 Pyruvic acid + 2 CoA → 2 acetyl − CoA + 2 CO2 + 4H

Krebs cycle:

2 acety1-CoA + 6H2 O + 2ADP − 2Pi → 4 CO2 + 2 ATP + 16 H + 2 CoA

Oxidative phosphorylation

24H + 6O2 + 34ADP + 34Pi → 12 H2O2 + 34 ATP

Glucose oxidation:

C6 H12 O6 + 6O2 + 38ADP + 38Pi → 6 CO2O + 6H2O + 38 ATP

The fate of pyruvic acid, and the method of disposal of hydrogen determines the type of respiration. In general, three types are recognized (Fig. 17-11).

Fermentation:

It is defined as a series of energy yielding reactions where an organic compound serves as hydrogen-acceptor.

This takes place in the absence of oxygen and two types are recognized: alcoholic fermentation and lactic acid fermentation (Fig. 17-11 A). While the former takes place in some bacteria, yeast and some green plants including germinating pollen under anaerobic conditions, the latter type is characteristic of animals and some bacteria.

In the alcoholic type, pyruvate is decarboxylated to acetaldehyde and the latter compound is reduced to ethyl alcohol by NADH. In the process NAD+ is released. Here acetaldehyde serves as hydrogen acceptor.

In the lactic acid fermentation, pyruvic acid formed acts as hydrogen acceptor and produces lactic acid and NAD+.

Both types of fermentation yield 2 molecules of ATP.