Glycolysis and the Citric Acid Cycle

Glycolysis represents an anabolic pathway common in both aerobic and anaerobic organisms. Other sugars and polysaccharides have to be transformed into glucose or one of its phosphorylated derivatives before being processed any further. In the course of degradation, ATP is produced. Pyruvate may be regarded as the preliminary final product of the degradation - in a strictly formal sense - because it is here that the pathway ramifies: pyruvate is hydrated under anaerobic conditions resulting in either lactate (in lactic-acid bacteria) or ethanol (in yeast). If glycolysis results in these final products, it is spoken of fermentation. Under aerobic conditions, pyruvate is fed into the citric acid cycle via an intermediate product. This pathway produces a neat amount of energy in the form of ATP. The starting product glucose is completely oxydized to water and carbon dioxide.

In 1905, A. HARDEN and W. YOUNG noticed, that the degradation of glucose stops if inorganic phosphate is not present in sufficient amounts. It is necessary to phosphorylate sugar. They were able to isolate a hexose diphosphate later on identified as fructose-1,6-diphosphate and could prove that is was an intermediate product of glucose degradation.

Glucose degradation was completely resolved in the years before 1940. The biochemists G. EMBDEN , O. MEYERHOFF, C. NEUBER, J. PARNASS, O. WARBURG and G. and C. CORI contributed most.

Glycolysis, sometimes also called the EMBDEN-MEYERHOFF-PARNASS-scheme consists of several reactions:

The pathway starts with an irreversible step, the ATP-consuming phosphorylation of glucose. In the second, reversible reaction glucose-6-phosphate is isomerized to fructose-6-phosphate.

Consumption of a second ATP leads to fructose-1,6-diphosphate. This step, too, is irreversible as the delta G° of the phosphorylated sugar is not sufficient to transfer the phosphate residue back to the ADP. These reactions are the only irreversible steps of all glycolysis. When drawing a linear structure of fructose-1,6-diphosphate ( = fructose-1,6-bisphosphate or FDP, it becomes clear that both phosphate groups are terminal so that the C-C linkage inbetween is weakened (dipoles at both ends of the molecule). This leads to an increased tendency of the molecule to disintegrate. With the aid of the enzyme aldolase, the fragments D-glyceraldehyde-3-phosphate (GAP) and dihydroxyacetone phosphate (DHAP) are thus produced. They can be interconversed since they are aldose-ketose isomers. Under physiological conditions far more dihydroxyacetone phosphate is present. But it is glyceraldehyde-3-phosphate that is continuously needed for subsequent reactions. It is steadily replaced by interconversion of dihydroxyacetone phosphates.

At this stage of glycolysis, the first oxidation ocurs though no oxygen is present. As a consequene of the dehydration of , NAD⁺ is reduced. Glyceraldhyde-3-phosphate is at the same time phosphorylated by addition of P_i.

The aldehyde group of glyceraldehyde-3-phosphate is thus transformed into a carboxy-group that is esterized to a phosphate. It is the exergonic aldehyde oxidation that drives the synthesis of the acyl phosphate 1,3-bisphosphoglycerate (1,3-BPG, also called 1,3-diphosphoglycerate). This molecule is very unstable and looses one of its phosphate groups easily to ADP. It is the first ATP-generating step of glycolysis. Two molecules of ATP had been invested and two are generated already during the first energy-producing step.

Why two? The original C₆ molecule has been transformed into two C₃ molecules so that we have to double both gains and losses from now on.

The two following steps are not that interesting but the dephosphorylation of phosphoenolpyruvate (PEP), the final common step of glycolysis is important again. The free energy of PEP hydrolysis is coupled to the synthesis of ATP to form pyruvate. The subsequent fate of pyruvate is, as has been mentioned above, dependend on the presence or absence of oxygen. Anaerobic processing, also called fermentation, leads to lactate or ethanol. Lactate is, for example, generated with the help of the enzyme lactate dehydrogenase and consumes NADH + H⁺. Yeast cells growing under anaerobic conditions produce ethanol with the aid of the enzymes pyruvate decarboxylase and alcohol dehydrogenase. They, too, need NADH + H⁺. The net gain of the fermentation process is rather low since no more ATP is produced. Aerobic conditions, in contrast, lead to the generation of acetyl-CoA:

pyruvate + NAD⁺ + CoA > acetyl-CoA + CO₂ + NADH + H⁺

The process is an oxidative decarboxylation since carbon dioxide is set free while pyruvate is dehydrated (oxydized). Thiamine pyrophosphate (TTP), the coenzyme of pyruvate dehydrogenase plays a decisive part in this reaction.

Acetyl-CoA C N O P S

© R. Bergmann

Citric Acid Cycle

Coenzyme A is a cofactor (a prosthetic group) like NAD⁺ and the others already discussed. The terminal SH-group is the active centre that binds to a C₂ unit (an acyl residue) and transfers it to another molecular group. Acetyl-CoA (activated acetic acid) is on one hand the starting compound of the fatty acid synthesis and on the other hand can the activated acetic acid be coupled to oxaloacetate. This coupling is the first reaction of the citric acid cycle.

Via a number of intermediate products, oxaloacetate is finally regenerated, the reason why this pathway is termed cycle.

But much more happens in its course:

1. In two steps of the pathway, carbon dioxide, one of the two oxidation products of glucose is set free. The C₂ molecule fed into the cycle is thus fully oxydized.

2. In four reactions, 2 H⁺ + 2 e^- are generated, in three of them are they transferred to NAD⁺, in one (in the dehydration of succinate to fumarate) to FAD.

3. Some of the intermediates are starting points of biosynthetic pathways.

4. One molecule of ATP (GTP in animals) is produced. Its generation is preceded by that of succinyl-CoA. This compound contains a CS-CoA-linkage (a thioester bond, delta G°: -8 kcal/mol = ca -33.6 kJ/mol). The breaking of this bond provides enough energy for the generation of one molecule of ATP (plants and bacteria) or GTP (mammals).