Gluconeogenesis

Gluconeogenesis (from the Greek "creation") is an anabolic metabolic pathway that allows the biosynthesis of glucose from non-glucidic precursors. It includes the use of various amino acids, lactate, pyruvate, glycerol and any of the intermediates of the tricarboxylic acid cycle (or Krebs cycle) as carbon sources for the metabolic pathway. All amino acids except leucine and lysine can supply carbon for glucose synthesis. Even-chain fatty acids do not provide carbons for glucose synthesis, since the result of their β-oxidation (Acetyl-CoA) is not a gluconeogenic substrate; while odd chain fatty acids will provide a carbon skeleton that will result in Acetyl-CoA and Succinyl-CoA (which is a gluconeogenic substrate as it is an intermediate of the Krebs cycle). Some tissues, such as the brain, erythrocytes, kidney, cornea of the eye and muscle, when the individual performs strenuous activity, require a continuous supply of glucose, obtaining it from glycogen from the liver, which can only satisfy these needs for 10 to 18 hours at most, which is how long it takes for the glycogen stored in the liver to be depleted. Subsequently, the formation of glucose begins from substrates other than glycogen.

Gluconeogenesis takes place almost exclusively in the liver (10% in the kidneys). It is a key process as it allows higher organisms to obtain glucose in metabolic states such as fasting.

Reactions of gluconeogenesis

The enzymes that participate in the glycolytic pathway also participate in gluconeogenesis; Both routes are differentiated by three irreversible reactions that use specific enzymes of this process and the two metabolic detours of this pathway.

These reactions are:

1. Glucose-6-phosphate to glucose.
2. fructose-1,6-bisphosphate to fructosa-6-phosphate.
3. From pyruvato to phosphoenolpiruvato.

Conversion of pyruvate to phosphoenolpyruvate

Oxaloacetate is an intermediate in the production of phosphoenolpyruvate in gluconeogenesis. The conversion of pyruvate to phosphoenolpyruvate in gluconeogenesis takes place in two steps. The first of these is the reaction of pyruvate and carbon dioxide to give oxaloacetate. This step requires energy, which is made available by hydrolysis of ATP.

The enzyme that catalyzes this reaction is pyruvate carboxylase, an allosteric enzyme found in the mitochondria. Acetyl-CoA is an allosteric effector that activates pyruvate carboxylase. When there is more acetyl-CoA than necessary to maintain the citric acid cycle, pyruvate is directed to gluconeogenesis. Magnesium ion and biotin are necessary for effective catalysis.

Biotin, covalently linked to the enzyme, reacts with CO₂, which is covalently linked. The CO₂ is then incorporated into pyruvate, thus forming oxaloacetate.

The conversion of oxaloacetate to phosphoenolpyruvate is catalyzed by the enzyme phosphoenolpyruvate carboxykinase, which is found in the mitochondria and cytosol. This reaction also includes the hydrolysis of a nucleoside triphosphate, in this case GTP instead of ATP.

Conversion of fructose-1,6-bisphosphate to fructose-6-phosphate

The phosphofructokinase 1 reaction of glycolysis is essentially irreversible, but only because it is driven by the transfer of phosphate from ATP. The reaction that takes place in gluconeogenesis to avoid this step consists of a simple hydrolytic reaction, catalyzed by fructose-1,6-bisphosphatase.

The multi-subunit enzyme requires the presence of Mg²⁺ for its activity and constitutes one of the main control sites that regulate the global gluconeogenesis pathway. The fructose-6-phosphate formed in this reaction subsequently undergoes isomerization to glucose-6-phosphate by the action of phosphoglucoisomerase.

Conversion of glucose-6-phosphate to glucose

Glucose-6-phosphate cannot be converted to glucose by the reverse action of hexokinase or glucokinase; The transfer of phosphate from ATP makes the reaction practically irreversible. Another enzyme specific to gluconeogenesis, glucose-6-phosphatase, which also requires Mg²⁺, takes action instead. This derivatization reaction also occurs by simple hydrolysis.

Glucose-6-phosphatase is found mainly in the endoplasmic reticulum of the liver with its active site on the luminal side (of the ER). The importance of its location in the liver is that a characteristic function of the liver is to synthesize glucose for export to tissues through blood circulation.

Regulation

The regulation of gluconeogenesis is crucial for many physiological functions, but above all for the proper functioning of nervous tissue. The flow through the pathway must increase or decrease, depending on the lactate produced by the muscles, glucose from the diet, or other gluconeogenic precursors.

Gluconeogenesis is largely controlled by diet. Animals that ingest abundant carbohydrates have low rates of gluconeogenesis, while fasting animals or those that ingest few carbohydrates have a high flux through this pathway.

Since gluconeogenesis synthesizes glucose and glycolysis catabolizes it, it is evident that gluconeogenesis and glycolysis must be controlled reciprocally. In other words, intracellular conditions that activate one pathway tend to inhibit the other.

Regulation by energy levels

Fructose 1,6-bisphosphatase is inhibited by high concentrations of AMP, associated with an energy-poor state. That is, the high concentration of AMP and reduced ATP inhibits gluconeogenesis

Regulation by fructose 2,6-bisphosphate

Fructose 1,6-bisphosphatase is inhibited by fructose 2,6-bisphosphate, an allosteric modulator whose concentration is determined by the circulating blood concentration of glucagon; Fructose 1,6-bisphosphatase is present in both the liver and kidneys.

Regulation of phosphorylation

This process is dependent on the concentration of ATP; As ATP concentration decreases, phosphorylation is also observed to decrease and vice versa. In the liver, this process increases by increasing the synthesis of glucokinase, a process that is promoted by insulin. The membrane of hepatocytes is very permeable to glucose; in muscle and adipose tissue, insulin acts on the membrane to make it permeable to it.

Allosteric regulation

Prolonged fasting increases acetyl-CoA and this stimulates pyruvate carboxylase and therefore gluconeogenesis, while inhibiting Pyruvate Dehydrogenase; Elevation of alanine and glutamine stimulate gluconeogenesis. Cortisol increases substrate availability and fructose 2,6-bisphosphate inhibits fructose 1,6-bisphosphatase.

Energy balance

We have highlighted that catabolic pathways generate energy, while anabolic pathways entail an energy cost. In the case of gluconeogenesis we can calculate this cost; Glucose synthesis is costly to the cell in an energetic sense. If we start from pyruvate, six high-energy phosphate groups are consumed: 4 ATP (due to the reactions of pyruvate carboxylase and phosphoglycerate kinase) and 2 GTP (a consequence of the decarboxylation of oxaloacetate), as well as 2 NADH, which is the energy equivalent of another 5 ATP (since mitochondrial oxidation of 1 NADH generates 2.5 ATP).

On the other hand, if glycolysis could act in the opposite direction, the energy expenditure would be much lower: 2 NADH and 2 ATP

Biomedical importance

Gluconeogenesis meets the body's needs for glucose when it is not available in sufficient quantities in the diet. A constant supply of glucose is required as a source of energy for the nervous system and erythrocytes. Furthermore, glucose is the only fuel that supplies energy to skeletal muscle under anaerobic conditions. Glucose is a precursor to milk sugar (lactose) in the mammary gland and is actively taken up by the baby. On the other hand, gluconeogenic mechanisms are used to purify the metabolism products of other tissues from the blood; for example, lactate, produced by muscle and erythrocytes, and glycerol, which is continuously formed by adipose tissue.