Tag Archives: glucose-alanine cycle

Glucose-alanine cycle

What is the glucose-alanine cycle?

The glucose-alanine cycle, or Cahill cycle, proposed for the first time by Mallette, Exton and Park, and Felig et al. between 1969 and 1970, consists of a series of steps through which extrahepatic tissues, for example the skeletal muscle, export pyruvate and amino groups as alanine to the liver, and  receive glucose from the liver via the bloodstream.
The main steps of the glucose-alanine cycle are summarized below.

  • When in extrahepatic tissues amino acids are used for energy, pyruvate, derived from the glycolytic pathway, is used as amino group acceptor, forming alanine, a nonessential amino acid.
  • Alanine diffuses into the bloodstream and reaches the liver.
  • In the liver, the amino group of alanine is transferred to α-ketoglutarate to form pyruvate and glutamate, respectively.
  • The amino group of glutamate mostly enters the urea cycle, and in part acts as a nitrogen donor in many biosynthetic pathways.
    Pyruvate enters the gluconeogenesis pathway and is used for glucose synthesis.
  • The newly formed glucose diffuses into the bloodstream and reaches the peripheral tissues where, due to glycolysis, is converted into pyruvate that can accept amino groups from the free amino acids, thus closing the cycle.

Therefore, the glucose-alanine cycle provides a link between carbohydrate and amino acid metabolism, as schematically described below.

Glucose → Pyruvate → Alanine → Pyruvate → Glucose

Glucose-Alanine Cycle
Fig. 1 – Glucose-Alanine Cycle

The glucose-alanine cycle occurs not only between the skeletal muscle, the first tissue in which it was observed, and the liver, but involves other cells and extrahepatic tissues including cells of the immune system, such as lymphoid organs.

The steps of the glucose-alanine cycle

The analysis of the steps of the glucose-alanine cycle is made considering the cycle between skeletal muscle and the liver.
Both intracellular and extracellular proteins are continuously hydrolyzed to the constituent amino acids and resynthesized, and the rate at which these processes occur is balanced precisely, thereby preventing loss of fat free mass.
However, under catabolic conditions, such as intense and prolonged exercise or fasting, the rate of muscle protein breakdown exceeds synthesis. This leads to the liberation of amino acids, some of which are used for energy and others for gluconeogenesis. And the oxidation of the carbon skeletons of amino acids, in particular branched chain amino acids or BCAA (leucine, isoleucine  and valine), may be a significant source of energy for the muscle. For example, after about 90 minutes of strenuous exercise, amino acid oxidation in muscle provides 10-15% of the energy needed for contraction.
The utilization of the carbon skeletons of amino acids for energy involves the removal of the amino group, and then the excretion of amino nitrogen in a non-toxic form.
The removal of the α-amino group occurs by transamination, that can be summarized as follows:

α-Keto acid + Amino acid ⇄ New amino acid + New α-keto acid

Such reactions, catalyzed by enzymes called aminotransferases or transaminases (EC 2.6.1) are freely reversible (see below).
Branched chain amino acids, for example, transfer the amino group to α-ketoglutarate or 2-oxoglutaric acid, to form glutamate and the α-keto acid derived from the original amino acid, in a reaction catalyzed by branched chain aminotransferase or BCAT (EC 2.6 .1.42).

The glucose-alanine cycle in skeletal muscle

In skeletal muscle, the newly formed glutamate may react with ammonia to form glutamine, for many tissues and organs, such as the brain, the major vehicle for interorgan transport of nitrogen. The reaction is catalyzed by the cytosolic enzyme glutamine synthetase (EC 6.3.1.2), and consumes an ATP.

Glutamate + NH4+ + ATP → Glutamine + ADP + Pi

In this case, glutamate leaves the Cahill cycle.
Alternatively, and in contrast to what happens in most of the other tissues, the newly formed glutamate may transfer the amino group to pyruvate, derived from glycolysis, to form alanine and α-ketoglutarate. This transamination is catalyzed by alanine aminotransferase or ALT (EC 2.6.1.2), an enzyme found in most animal and plant tissues.

Glutamate + Pyruvate ⇄ Alanine + α-Ketoglutarate

The alanine produced and that derived directly from protein breakdown, and muscle proteins are rich in alanine, can leave the cell and be carried by the bloodstream to the liver; in this way the amino group reaches the liver. And the rate at which alanine formed by transamination of pyruvate is transferred into the circulation is proportional to the intracellular pyruvate production.
Note: alanine and glutamine are the major sources of nitrogen and carbon in interorgan amino acid metabolism.

The glucose-alanine cycle in the liver

Once in the liver, a hepatic alanine aminotransferase catalyzes a transamination in which alanine, the major gluconeogenic amino acid, acts as an amino group donor and α-ketoglutarate as an α-keto acid acceptor. The products of the reaction are pyruvate, i.e. the carbon skeleton of alanine, and glutamate.

Alanine + α-Ketoglutarate ⇄ Glutamate + Pyruvate

Glutamate, in the reaction catalyzed by glutamate dehydrogenase (EC 1.4.1.2), an enzyme present in the mitochondrial matrix, forms ammonium ion, which enters the urea cycle, and α-ketoglutarate, which can enter the Krebs cycle. This reaction is an anaplerotic reaction that links amino acid metabolism with the Krebs cycle.

Glucose-Alanine Cycle

However, glutamate can also react  with oxaloacetate to form aspartate and α-ketoglutarate, in a reaction catalyzed by aspartate aminotransferase (EC 2.6.1.1). Aspartate is involved in the formation of urea as well as in the synthesis of purines and pyrimidines.

Glutamate + Oxaloacetate ⇄ Aspartate + α-Ketoglutarate

Also the pyruvate produced may have different metabolic fates: it can be oxidized for ATP production, and then leave the glucose-alanine cycle, or enter the gluconeogenesis pathway, and thus continue in the cycle.
The glucose produced is released from the liver into the bloodstream and delivered to various tissues that require it, as the skeletal muscle, in which it is used for pyruvate synthesis. In turn, the newly formed pyruvate may react with glutamate, thus closing the cycle.

Transaminases

As previously mentioned, the removal of the amino group from amino acids occurs through transamination (see above for the general reaction). These reactions are catalyzed by enzymes called aminotransferases or transaminases.
They are cytosolic enzymes, present in all cells and particularly abundant in the liver, kidney, intestine and muscle; they require pyridoxal phosphate or PLP, the active form of vitamin B6 or pyridoxine, as a coenzyme, which is tightly bound to the active site.
In transamination reactions, the amino group of free amino acids, except of threonine and lysine, is channeled towards a small number of α-keto acids, notably pyruvate, oxaloacetate and α-ketoglutarate.
Cells contain different types of aminotransferases: many are specific for α-ketoglutarate as α-keto acid acceptor, but differ in specificity for the amino acid, from which they are named. Examples are the aforementioned alanine aminotransferase, also called alanine transaminase and glutamic pyruvic transferase or GPT, and aspartate aminotransferase or AST, also called glutamic-oxaloacetic transaminase or GOT.
It should be underlined that there is no net deamination in these reactions, no loss of amino groups, as the α-keto acid acceptor is aminated and the amino acid deaminated.

Functions of the glucose-alanine cycle

This cycle has various functions.

  • It transports nitrogen in a non-toxic form from peripheral tissues to the liver.
  • It transports pyruvate, a gluconeogenic substrate, to the liver.
  • It removes pyruvate from peripheral tissues.  This leads to a higher production of ATP from glucose in these tissues. In fact, the NADH produced during glycolysis can enter the mitochondria and be oxidized through oxidative phosphorylation.
  • It allows to maintain a relatively high concentration of alanine in hepatocytes, sufficient to inhibit protein degradation.
  • It may play a role in host defense against infectious diseases.

Finally, it is important to underline that there is no net synthesis of glucose in the glucose-alanine cycle.

Energy cost of the glucose-alanine cycle

Like the Cori cycle, also the glucose-alanine cycle has an energy cost, equal to 3-5 ATP.
The part of the cycle that takes place in peripheral tissues involves the production of 5-7 ATP per molecule of glucose:

  • 2 ATP are produced by glycolysis;
  • 3-5 ATP derive from NADH/FADH2 (see below).

Instead in the liver, gluconeogenesis and the urea cycle cost 10 ATP:

  • 6 ATP are consumed in the during gluconeogenesis per molecule of glucose synthesized;
  • 4 ATP are consumed in the urea cycle per molecule of urea synthesized.

The glucose-alanine cycle, like the Cori cycle, shifts part of the metabolic burden from extrahepatic tissues to the liver. However, the energy cost paid by the liver is justified by the advantages that the cycle brings to the whole body, as it allows, in particular conditions, an efficient breakdown of proteins in extrahepatic tissues (especially skeletal muscle), which in turn allows to obtain gluconeogenic substrates as well as the use of amino acids for energy in extrahepatic tissues.

Similarities and differences between glucose-alanine cycle and Cori cycle?

There are some analogies between the two cycles, which are listed below.

  • The Cahill cycle partially overlaps the Cori cycle when pyruvate is converted to glucose and the monosaccharide is transported to extrahepatic tissues, in which it is converted again to pyruvate via the glycolytic pathway.
  • The entry into gluconeogenesis pathway is similar for the two cycles: both alanine and lactate are converted to pyruvate.
  • Like the Cori cycle, the glucose-alanine cycle occurs between different cell types, unlike metabolic pathways such as glycolysis, Krebs cycle or gluconeogenesis that occur within individual cells
Glucose-Alanine Cycle
Fig. 2 – Glucose-Alanine Cycle and Cori Cycle

Below, some differences between the two cycles.

  • The main difference concerns the three carbon intermediate that from peripheral tissues reach the liver: lactate in the Cori cycle, and alanine in the glucose-alanine cycle.
  • Another difference concerns the fate of the NADH produced by glycolysis in peripheral tissues.
    In the Cori cycle, the coenzyme acts as reducing agent to reduce pyruvate to lactate, in the reaction catalyzed by lactate dehydrogenase (EC 1.1.1.27).
    In the glucose-alanine cycle, this reduction does not occur and the electrons of NADH can be transported into the mitochondria via the malate-aspartate and glycerol 3-phosphate shuttles, generating NADH, the first shuttle, and FADH2, the other shuttle. And the yield of ATP from NADH and FADH2 is 2.5 and 1.5, respectively.
  • Finally, from the previous point, it is clear that, unlike the Cori cycle, the Cahill cycle requires the presence of oxygen and mitochondria in the peripheral tissues.
References

Berg J.M., Tymoczko J.L., and Stryer L. Biochemistry. 5th Edition. W. H. Freeman and Company, 2002

Felig P., Pozefsk T., Marlis E., Cahill G.F. Alanine: key role in gluconeogenesis. Science 1970;167(3920):1003-4 [Abstract]

Gropper S.S., Smith J.L., Groff J.L. Advanced nutrition and human metabolism. Cengage Learning, 2009 [Google eBooks]

Lecker S.H., Goldberg A.L. and Mitch W.E. Protein degradation by the ubiquitin–proteasome pathway in normal and disease states. J Am Soc Nephrol 2006;17(7):1807-19 [PDF]

Mallette L. E., Exton J. H., and Park C. R. Control of gluconeogenesis from amino acids in the perfused  rat liver. J Biol Chem 1969;244(20):5713-23 [PDF]

Nelson D.L., M. M. Cox M.M. Lehninger. Principles of biochemistry. 4th Edition. W.H. Freeman and Company, 2004

Raju S.M., Madala B. Illustrated medical biochemistry. Jaypee Brothers Publishers, 2005 [Google eBooks]

Wu G. Amino acids: biochemistry and nutrition. CRC Press, 2013 [Google eBooks]

Cori cycle: definition, function, biochemistry, involved tissues

What is the Cori cycle?

The Cori cycle, or glucose-lactate cycle, was discovered by Carl Ferdinand Cori and Gerty Theresa Radnitz, a husband-and-wife team, in the ‘30s and ‘40s of the last century . They demonstrated the existence of a metabolic cooperation between the skeletal muscle working under low oxygen conditions and the liver. This cycle can be summarized as follows:

  • the conversion of glucose to lactic acid, or lactate, by anaerobic glycolysis in skeletal muscle cells;
  • the diffusion of lactate from muscle cells into the bloodstream, by which it is transported to the liver;
  • the conversion of lactate to glucose by hepatic gluconeogenesis;
  • the diffusion of glucose from the hepatocytes into the bloodstream, by which it is transported back to the skeletal muscle cells, thereby closing the cycle.

Summarizing, we have: part of the lactate produced in skeletal muscle is converted to glucose in the liver, and transported back to skeletal muscle, thus closing the cycle.

Glucose → Lactate →Glucose

The importance of this cycle is demonstrated by the fact that it may account for about 40% of plasma glucose turnover.

Where does the Cori cycle occur?

In addition to skeletal muscle, this metabolic cooperation was also demonstrated between other extrahepatic tissues and liver.  Indeed, like the glucose-alanine cycle, the glucose-lactate cycle is active between the liver and all those tissues that do not completely oxidize glucose to CO2 and H2O, in which case pyruvate for conversion to lactate or, by transamination, to alanine would lack (see below).
In addition to skeletal muscle cells, examples of cells that continually produce lactic acid are red blood cells, immune cells in the lymph nodules,  proliferating cells in the bone marrow, and epithelial cells in the skin.
Note: skeletal muscle produces lactic acid even at rest, although at low rate.

Cori Cycle
Fig. 1 – The Cori Cycle

From a biochemical point of view, the Cori cycle links gluconeogenesis with anaerobic glycolysis, using different tissues to compartmentalize opposing metabolic pathways. In fact, in the same cell, regardless of the cell type, these metabolic pathways are not very active simultaneously. Glycolysis is more active when the cell requires ATP; by contrast, when the demand for ATP is low, gluconeogenesis, in those cells where it occurs, is more active.
And it is noteworthy that, although traditionally the metabolic pathways, such as glycolysis, citric acid cycle, or gluconeogenesis, are considered to be confined within individual cells, the Cori cycle, as well as the glucose-alanine cycle, occurs between different cell types.
Finally, it should be underscored that the Cori cycle also involves the renal cortex, particularly the proximal tubules, another site where gluconeogenesis occurs.

The steps of the Cori cycle

The analysis of the steps of the Cori cycle is made considering the lactate produced by red blood cells and skeletal muscle cells.
Mature red blood cells are devoid of mitochondria, nucleus and ribosomes, and obtain the necessary energy only by glycolysis. The availability of NAD+ is essential for glycolysis to proceed as well as for its rate: the oxidized form of the coenzyme is required for the oxidation of glyceraldehyde 3-phosphate to 1,3-bisphosphoglycerate in the reaction catalyzed by glyceraldehyde 3-phosphate dehydrogenase (EC 1.2.1.12).

Glyceraldehyde 3-phosphate + NAD+ → 1,3-Bisphosphoglycerate + NADH + H+

The accumulation of NADH is avoided by the reduction of pyruvate to lactate, in the reaction catalyzed by lactate dehydrogenase (EC 1.1.1.27), where NADH acts as reducing agent.

Pyruvate + NADH + H+ → Lactate + NAD+

The skeletal muscle, particularly fast-twitch fibers which contain a reduced number of mitochondria, under low oxygen condition, such as during intense exercise, produces significant amounts of lactate. In fact, in such conditions:

  • the rate of pyruvate production by glycolysis  exceeds the rate of its oxidation by the citric acid cycle, so that less than 10% of the pyruvate enters the citric acid cycle;
  • the rate at which oxygen is taken up by the cells is not sufficient to allow aerobic oxidation of all the NADH  produced.

And, like in red blood cells, the reaction catalyzed by lactate dehydrogenase, regenerating NAD+, allows glycolysis to proceed.
However, lactate is an end product of metabolism that must be converted back into pyruvate to be used.
The plasma membrane of most cells is freely permeable to both pyruvate and lactate that can thus reach the bloodstream. And, regarding for example the skeletal muscle, the amount of lactate that leaves the cell is greater than that of pyruvate due to the high NADH/NAD+ ratio in the cytosol and to the catalytic properties of the skeletal muscle isoenzyme of LDH.
Once into the bloodstream, lactate reaches the liver, which is its major user, where it is oxidized to pyruvate in the reaction catalyzed by the liver isoenzyme of lactate dehydrogenase (see below).

Lactate + NAD+ → Pyruvate + NADH + H+

In the hepatocyte, this oxidation is favored by the low NADH/NAD+ ratio in the cytosol.
Then, pyruvate enters the gluconeogenesis pathway to be converted into glucose.
Glucose leaves the liver, enters into the bloodstream and is delivered to the muscle, as well as to other tissues and cells that require it, such as red blood cells and neurons, thus closing the cycle.

Lactate dehydrogenase

The enzyme is a tetramer composed of two different types of subunits, designed as:

  • H subunit (heart) or B chain;
  • M subunit (muscle) or A chain.

The H subunit predominates in the heart, whereas the M subunit predominates in the  skeletal muscle and liver. Typically, tissues in which a predominantly or exclusively aerobic metabolism occurs, such as the heart, synthesize H subunits to a greater extent than M subunits, whereas tissues in which anaerobic metabolism is important, such as skeletal muscle, synthesize M subunits to a greater extent than H subunits.
The two subunits associate in 5 different ways to form homopolymers, that is, macromolecules formed by repeated, identical subunits, or heteropolymers, that is, macromolecules formed by different subunits. Different LDH  isoenzymes have different catalytic properties, as well as different distribution in various tissues, as indicated below:

  • H4, also called type 1, LDH1, or A4, a homopolymer of H subunits, is found in cardiac muscle, kidney, and red blood cells;
  • H3M1, also called type 2, LDH2, or A3B, has a tissue distribution similar to that of LDH1;
  • H2M2, also called type 3, LDH3, or A2B2, is found in the spleen, brain, white cells, kidney, and lung;
  • H1M3, also called type 4, LDH4, or AB3, is found in the spleen, lung, skeletal muscle, lung, red blood cells, and kidney;
  • M4, also called type 5, LDH5, or B4, a homopolymer of M subunits, is found in the liver, skeletal muscle, and spleen.

The H4 isoenzyme has a higher substrate affinity than the M4 isoenzyme.
The H4 isoenzyme is allosterically inhibited by high levels of pyruvate (its product), whereas the M4 isoenzyme is not.
The other LDH isoenzymes have intermediate properties, depending on the ratio between the two types of subunits.
It is thought that the H4 isoenzyme is the most suitable for catalyzing the oxidation of lactate to pyruvate that, in the heart, due to its exclusively aerobic metabolism, is then completely oxidized to CO2 and H2O. Instead, the M4 isoenzyme is the main isoenzyme found in skeletal muscle, most suitable for catalyzing the reduction of pyruvate to lactate, thus allowing glycolysis to proceed in anaerobic conditions.

Other metabolic fates of lactate

From the above, it is clear that lactate is not a metabolic dead end, a waste product of glucose metabolism.
And it may have a different fate from that entering the Cori cycle.
For example, in skeletal muscle during recovery following an exhaustive exercise, that is, when oxygen is again available, or if the exercise is of low intensity, lactate is re-oxidized to pyruvate, due to NAD+ availability, and then completely oxidized to CO2 and H20, with a greater production of ATP than in anaerobic condition. In such conditions, the energy stored in NADH will be released, yielding on average 2.5 ATP per molecule of NADH.
In addition, lactate can be taken up by exclusively aerobic tissues, such as heart, to be oxidized to CO2 and H20.

Energy cost of the Cori cycle

The Cori cycle results in a net consumption of 4 ATP.
The gluconeogenic leg of the cycle consumes 2 GTP and 4 ATP per molecule of glucose synthesized, that is, 6 ATP.
The ATP-consuming reactions are catalyzed by:

  • pyruvate carboxylase (EC 6.4.1.1): an ATP;
  • phosphoenolpyruvate carboxykinase (EC 4.1.1.32): a GTP;
  • glyceraldehyde 3-phosphate dehydrogenase (EC 1.2.1.12): an ATP.

Since two molecules of lactate are required for the synthesis of one molecule of glucose, the net cost is 2×3=6 high energy bonds per molecule of glucose.
Conversely, the glycolytic leg of the cycle produces only 2 ATP per molecule of glucose.
Therefore, more energy is required to produce glucose from lactate than that obtained by anaerobic glycolysis in extrahepatic tissues. This explains why the Cori cycle cannot be sustained indefinitely.

Is the Cori cycle a futile cycle?

The continuous breakdown and resynthesis of glucose, feature of the Cori cycle, might seem like a waste of energy. Indeed, this cycle allows the effective functioning of many extrahepatic cells at the expense of the liver and partly of the renal cortex. Below, two examples.

  • Red blood cells
    These cells, lacking a nucleus, ribosomes, and mitochondria, are smaller than most other cells. Their small size allows them to pass through tiny capillaries. However, the lack of mitochondria makes them completely dependent on anaerobic glycolysis for ATP production. Then, the lactate is partly disposed of by the liver and renal cortex.
  • Skeletal muscle
    Its cells, and particularly fast-twitch fibers contracting under low oxygen conditions, such as during intense exercise, produce much lactate.
    In such conditions, anaerobic glycolysis leads to the production of 2 ATP per molecule of glucose, 3 if the glucose comes from muscle glycogen, therefore, much lower than the 29-30 ATP produced by the complete oxidation of the monosaccharide. However, the rate of ATP production by anaerobic glycolysis is greater than that produced by the complete oxidation of glucose. Therefore, to meet the energy requirements of contracting muscle, anaerobic glycolysis is an effective means of ATP production. But this could lead to an intracellular accumulation of lactate, and a consequent reduction in intracellular pH. Obviously, such accumulation does not occur, due also to the Cori cycle, in which the liver pays the cost of the disposal of a large part of the muscle lactate, thereby allowing the muscle to use ATP for the contraction.
    And the oxygen debt, which always occurs after a strenuous exercise, is largely due to the increased oxygen demand of the hepatocytes, in which the oxidation of fatty acids, their main fuel, provides the ATP required for gluconeogenesis from lactate.
  • During trauma, sepsis, burns, or after major surgery, an intense cell proliferation occurs in the wound, that is a hypoxic tissue, and in bone marrow. This in turn results in greater production of lactic acid, an increase in the flux through the Cori cycle and an increase in ATP consumption in the liver, which, as previously said, is supported by an increase in fatty acid oxidation. Hence, the nutrition plan provided to these patients must be taken into account this increase in energy consumption.
  • A similar condition seems to occur also in cancer patients with progressive weight loss.
  • The Cori cycle is also important during overnight fasting and starvation.

The Cori cycle and glucose-alanine cycle

These cycles are metabolic pathways that contribute to ensure a continuous delivery of glucose to tissues for which the monosaccharide is  the primary source of energy.
The main difference between the two cycles consists in the three carbon intermediate which is recycled: in the Cori cycle, carbon returns to the liver in the form of pyruvate, whereas in the glucose-alanine cycle in the form of alanine.
For more information, see: glucose-alanine cycle.

References

Bender D.A. Introduction to nutrition and metabolism. 3rd Edition. Taylor & Francis, 2004

Berg J.M., Tymoczko J.L., and Stryer L. Biochemistry. 5th Edition. W. H. Freeman and Company, 2002

Iqbal S.A., Mido Y. Biochemistry. Discovery Publishing House, 2005 [Google eBook]

Nelson D.L., M. M. Cox M.M. Lehninger. Principles of biochemistry. 4th Edition. W.H. Freeman and Company, 2004

Newsholme E.A., Leech T.R. Functional biochemistry in health and disease. John Wiley J. & Sons, Inc., Publication, 2010 [Google eBook]

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Rosenthal M.D., Glew R.H. Medical biochemistry – Human metabolism in health and disease. John Wiley J. & Sons, Inc., Publication, 2009

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