Themetabolic pathway of glycolysis convertsglucose topyruvate via a series of intermediate metabolites. Each chemical modification is performed by a different enzyme. Steps 1 and 3 consumeATP and steps 7 and 10 produce ATP. Since steps 6–10 occur twice per glucose molecule, this leads to a net production of ATP.
Summary of the 10 reactions of the glycolysis pathway
The wide occurrence of glycolysis in other species indicates that it is an ancient metabolic pathway.[2] Indeed, the reactions that make up glycolysis and its parallel pathway, thepentose phosphate pathway, can occur in theoxygen-free conditions of theArchean oceans, also in the absence of enzymes, catalyzed by metal ions, meaning this is a plausible prebiotic pathway forabiogenesis.[3]
The most common type of glycolysis is theEmbden–Meyerhof–Parnas (EMP) pathway, which was discovered byGustav Embden,Otto Meyerhof, andJakub Karol Parnas. Glycolysis also refers to other pathways, such as theEntner–Doudoroff pathway and various heterofermentative and homofermentative pathways. However, the discussion here will be limited to the Embden–Meyerhof–Parnas pathway.[4]
The glycolysis pathway can be separated into two phases:[5]
Investment phase – wherein ATP is consumed
Yield phase – wherein more ATP is produced than originally consumed
The use of symbols in this equation makes it appear unbalanced with respect to oxygen atoms, hydrogen atoms, and charges. Atom balance is maintained by the two phosphate (Pi) groups:[6]
Each exists in the form of ahydrogen phosphate anion ([HPO4]2−), dissociating to contribute2H+ overall
Each liberates an oxygen atom when it binds to anadenosine diphosphate (ADP) molecule, contributing 2O overall
Charges are balanced by the difference between ADP and ATP. In the cellular environment, all three hydroxyl groups of ADP dissociate into −O− and H+, giving ADP3−, and this ion tends to exist in an ionic bond with Mg2+, giving ADPMg−. ATP behaves identically except that it has four hydroxyl groups, giving ATPMg2−. When these differences along with the true charges on the two phosphate groups are considered together, the net charges of −4 on each side are balanced.[citation needed]
In high-oxygen (aerobic) conditions, eukaryotic cells can continue from glycolysis to metabolise the pyruvate through thecitric acid cycle or theelectron transport chain to produce significantly more ATP.
Importantly, under low-oxygen (anaerobic) conditions, glycolysis is the only biochemical pathway in eukaryotes that can generate ATP, and, for many anaerobic respiring organisms the most important producer of ATP.[7] Therefore, many organisms have evolvedfermentation pathways to recycle NAD+ to continue glycolysis to produce ATP for survival. These pathways includeethanol fermentation andlactic acid fermentation.
The modern understanding of the pathway of glycolysis took almost 100 years to fully learn.[8] The combined results of many smaller experiments were required to understand the entire pathway.
The first steps in understanding glycolysis began in the 19th century. For economic reasons, the French wine industry sought to investigate why wine sometimes turned distasteful, instead of fermenting into alcohol. The French scientistLouis Pasteur researched this issue during the 1850s.[9] His experiments showed that alcohol fermentation occurs by the action of livingmicroorganisms, yeasts, and that glucose consumption decreased under aerobic conditions (thePasteur effect).[10]
Eduard Buchner discovered cell-free fermentation.
The component steps of glycolysis were first analysed by the non-cellular fermentation experiments ofEduard Buchner during the 1890s.[11][12] Buchner demonstrated that the conversion of glucose to ethanol was possible using a non-living extract of yeast, due to the action ofenzymes in the extract.[13]: 135–148 This experiment not only revolutionized biochemistry, but also allowed later scientists to analyze this pathway in a more controlled laboratory setting. In a series of experiments (1905–1911), scientistsArthur Harden andWilliam Young discovered more pieces of glycolysis.[14] They discovered the regulatory effects of ATP on glucose consumption during alcohol fermentation. They also shed light on the role of one compound as a glycolysis intermediate: fructose 1,6-bisphosphate.[13]: 151–158
The elucidation of fructose 1,6-bisphosphate was accomplished by measuringCO2 levels when yeast juice was incubated with glucose.CO2 production increased rapidly then slowed down. Harden and Young noted that this process would restart if an inorganic phosphate (Pi) was added to the mixture. Harden and Young deduced that this process produced organic phosphate esters, and further experiments allowed them to extract fructose diphosphate (F-1,6-DP).
Arthur Harden andWilliam Young along with Nick Sheppard determined, in a second experiment, that a heat-sensitive high-molecular-weight subcellular fraction (the enzymes) and a heat-insensitive low-molecular-weight cytoplasm fraction (ADP, ATP and NAD+ and othercofactors) are required together for fermentation to proceed. This experiment begun by observing that dialyzed (purified) yeast juice could not ferment or even create a sugar phosphate. This mixture was rescued with the addition of undialyzed yeast extract that had been boiled. Boiling the yeast extract renders all proteins inactive (as it denatures them). The ability of boiled extract plus dialyzed juice to complete fermentation suggests that the cofactors were non-protein in character.[14]
Otto Meyerhof, one of the main scientists involved in completing the puzzle of glycolysis
In the 1920sOtto Meyerhof was able to link together some of the many individual pieces of glycolysis discovered by Buchner, Harden, and Young. Meyerhof and his team were able to extract different glycolytic enzymes frommuscle tissue, and combine them to artificially create the pathway from glycogen to lactic acid.[15][16]
In one paper, Meyerhof and scientist Renate Junowicz-Kockolaty investigated the reaction that splits fructose 1,6-diphosphate into the two triose phosphates. Previous work proposed that the split occurred via 1,3-diphosphoglyceraldehyde plus an oxidizing enzyme and cozymase. Meyerhoff and Junowicz found that the equilibrium constant for the isomerase and aldoses reaction were not affected by inorganic phosphates or any other cozymase or oxidizing enzymes. They further removed diphosphoglyceraldehyde as a possible intermediate in glycolysis.[16]
With all of these pieces available by the 1930s,Gustav Embden proposed a detailed, step-by-step outline of that pathway we now know as glycolysis.[17] The biggest difficulties in determining the intricacies of the pathway were due to the very short lifetime and low steady-state concentrations of the intermediates of the fast glycolytic reactions. By the 1940s, Meyerhof, Embden and many other biochemists had finally completed the puzzle of glycolysis.[16] The understanding of the isolated pathway has been expanded in the subsequent decades, to include further details of its regulation and integration with other metabolic pathways.
The first five steps of Glycolysis are regarded as the preparatory (or investment) phase, since they consume energy to convert the glucose into two three-carbon sugar phosphates[5] (G3P).
Once glucose enters the cell, the first step is phosphorylation of glucose by a family of enzymes calledhexokinases to form glucose 6-phosphate (G6P). This reaction consumes ATP, but it acts to keep the glucose concentration inside the cell low, promoting continuous transport of blood glucose into the cell through the plasma membrane transporters. In addition, phosphorylation blocks the glucose from leaking out – the cell lacks transporters for G6P, and free diffusion out of the cell is prevented due to the charged nature of G6P. Glucose may alternatively be formed from thephosphorolysis orhydrolysis of intracellular starch or glycogen.
Inanimals, anisozyme of hexokinase calledglucokinase is also used in the liver, which has a much lower affinity for glucose (Km in the vicinity of normal glycemia), and differs in regulatory properties. The different substrate affinity and alternate regulation of this enzyme are a reflection of the role of the liver in maintaining blood sugar levels.
The change in structure is an isomerization, in which the G6P has been converted to F6P. The reaction requires an enzyme, phosphoglucose isomerase, to proceed. This reaction is freely reversible under normal cell conditions. However, it is often driven forward because of a low concentration of F6P, which is constantly consumed during the next step of glycolysis. Under conditions of high F6P concentration, this reaction readily runs in reverse. This phenomenon can be explained throughLe Chatelier's Principle. Isomerization to a keto sugar is necessary for carbanion stabilization in the fourth reaction step (below).
The energy expenditure of another ATP in this step is justified in 2 ways: The glycolytic process (up to this step) becomes irreversible, and the energy supplied destabilizes the molecule. Because the reaction catalyzed byphosphofructokinase 1 (PFK-1) is coupled to the hydrolysis of ATP (an energetically favorable step) it is, in essence, irreversible, and a different pathway must be used to do the reverse conversion duringgluconeogenesis. This makes the reaction a key regulatory point (see below).
Furthermore, the second phosphorylation event is necessary to allow the formation of two charged groups (rather than only one) in the subsequent step of glycolysis, ensuring the prevention of free diffusion of substrates out of the cell.
The same reaction can also be catalyzed bypyrophosphate-dependent phosphofructokinase (PFP orPPi-PFK), which is found in most plants, some bacteria, archea, and protists, but not in animals. This enzyme uses pyrophosphate (PPi) as a phosphate donor instead of ATP. It is a reversible reaction, increasing the flexibility of glycolytic metabolism.[18] A rarer ADP-dependent PFK enzyme variant has been identified in archaean species.[19]
Destabilizing the molecule in the previous reaction allows the hexose ring to be split byaldolase into two triose sugars:dihydroxyacetone phosphate (a ketose), andglyceraldehyde 3-phosphate (an aldose). There are two classes of aldolases: class I aldolases, present in animals and plants, and class II aldolases, present in fungi and bacteria; the two classes use different mechanisms in cleaving the ketose ring.
Electrons delocalized in the carbon-carbon bond cleavage associate with the alcohol group. The resulting carbanion is stabilized by the structure of the carbanion itself via resonance charge distribution and by the presence of a charged ion prosthetic group.
Triosephosphate isomerase rapidly interconverts dihydroxyacetone phosphate withglyceraldehyde 3-phosphate (GADP) that proceeds further into glycolysis. This is advantageous, as it directs dihydroxyacetone phosphate down the same pathway as glyceraldehyde 3-phosphate, simplifying regulation.
The second half of glycolysis is known as the pay-off phase, characterised by a net gain of the energy-rich molecules ATP and NADH.[5] Since glucose leads to two triose sugars in the preparatory phase, each reaction in the pay-off phase occurs twice per glucose molecule. This yields 2 NADH molecules and 4 ATP molecules, leading to a net gain of 2 NADH molecules and 2 ATP molecules from the glycolytic pathway per glucose.
The hydrogen is used to reduce two molecules ofNAD+, a hydrogen carrier, to give NADH+ H+ for each triose.
Hydrogen atom balance and charge balance are both maintained because the phosphate (Pi) group actually exists in the form of ahydrogen phosphate anion (HPO2−4),[6] which dissociates to contribute the extra H+ ion and gives a net charge of -3 on both sides.
Here,arsenate ([AsO4]3−), an anion akin to inorganic phosphate may replace phosphate as a substrate to form 1-arseno-3-phosphoglycerate. This, however, is unstable and readily hydrolyzes to form3-phosphoglycerate, the intermediate in the next step of the pathway. As a consequence of bypassing this step, the molecule of ATP generated from1-3 bisphosphoglycerate in the next reaction will not be made, even though the reaction proceeds. As a result, arsenate is an uncoupler of glycolysis.[20]
This step is the enzymatic transfer of a phosphate group from1,3-bisphosphoglycerate to ADP byphosphoglycerate kinase, forming ATP and3-phosphoglycerate. At this step, glycolysis has reached the break-even point: 2 molecules of ATP were consumed, and 2 new molecules have now been synthesized. This step, one of the twosubstrate-level phosphorylation steps, requires ADP; thus, when the cell has plenty of ATP (and little ADP), this reaction does not occur. Because ATP decays relatively quickly when it is not metabolized, this is an important regulatory point in the glycolytic pathway.
ADP actually exists as ADPMg−, and ATP as ATPMg2−, balancing the charges at −5 both sides.
Cofactors: 2 Mg2+, one "conformational" ion to coordinate with the carboxylate group of the substrate, and one "catalytic" ion that participates in the dehydration.
A finalsubstrate-level phosphorylation now forms a molecule ofpyruvate and a molecule of ATP by means of the enzymepyruvate kinase. This serves as an additional regulatory step, similar to the phosphoglycerate kinase step.
The existence of more than one point of regulation indicates that intermediates between those points enter and leave the glycolysis pathway by other processes. For example, in the first regulated step,hexokinase converts glucose into glucose-6-phosphate. Instead of continuing through the glycolysis pathway, this intermediate can be converted into glucose storage molecules, such asglycogen orstarch. The reverse reaction, breaking down, e.g., glycogen, produces mainly glucose-6-phosphate; very little free glucose is formed in the reaction. The glucose-6-phosphate so produced can enter glycolysisafter the first control point.
In the second regulated step (the third step of glycolysis),phosphofructokinase converts fructose-6-phosphate into fructose-1,6-bisphosphate, which then is converted into glyceraldehyde-3-phosphate and dihydroxyacetone phosphate. The dihydroxyacetone phosphate can be removed from glycolysis by conversion into glycerol-3-phosphate, which can be used to form triglycerides.[21] Conversely,triglycerides can be broken down into fatty acids and glycerol; the latter, in turn, can beconverted into dihydroxyacetone phosphate, which can enter glycolysisafter the second control point.
The change in free energy, ΔG, for each step in the glycolysis pathway can be calculated using ΔG = ΔG°′ +RTlnQ, whereQ is thereaction quotient. This requires knowing the concentrations of themetabolites. All of these values are available forerythrocytes, with the exception of the concentrations of NAD+ and NADH. The ratio ofNAD+ to NADH in the cytoplasm is approximately 1000, which makes the oxidation of glyceraldehyde-3-phosphate (step 6) more favourable.
Using the measured concentrations of each step, and the standard free energy changes, the actual free energy change can be calculated. (Neglecting this is very common—the delta G of ATP hydrolysis in cells is not the standard free energy change of ATP hydrolysis quoted in textbooks).
Change in free energy for each step of glycolysis[22]: 582–583
From measuring the physiological concentrations of metabolites in an erythrocyte it seems that about seven of the steps in glycolysis are in equilibrium for that cell type. Three of the steps—the ones with large negative free energy changes—are not in equilibrium and are referred to asirreversible; such steps are often subject to regulation.
Step 5 in the figure is shown behind the other steps, because that step is a side-reaction that can decrease or increase the concentration of the intermediate glyceraldehyde-3-phosphate. That compound is converted to dihydroxyacetone phosphate by the enzyme triose phosphate isomerase, which is acatalytically perfect enzyme; its rate is so fast that the reaction can be assumed to be in equilibrium. The fact that ΔG is not zero indicates that the actual concentrations in the erythrocyte are not accurately known.
The enzymes that catalyse glycolysis are regulated via a range of biological mechanisms in order to control overallflux though the pathway. This is vital for bothhomeostatsis in a static environment, andmetabolic adaptation to a changing environment or need.[23] The details of regulation for some enzymes are highly conserved between species, whereas others vary widely.[24][25]
Allosteric inhibition and activation byProtein-protein interactions (PPI).[29] Indeed, some proteins interact with and regulate multiple glycolytic enzymes.[30]
In animals, regulation of blood glucose levels by the pancreas in conjunction with the liver is a vital part ofhomeostasis. Thebeta cells in thepancreatic islets are sensitive to the blood glucose concentration.[32] A rise in the blood glucose concentration causes them to releaseinsulin into the blood, which has an effect particularly on the liver, but also onfat andmuscle cells, causing these tissues to remove glucose from the blood. When the blood sugar falls the pancreatic beta cells cease insulin production, but, instead, stimulate the neighboring pancreaticalpha cells to releaseglucagon into the blood.[32] This, in turn, causes the liver to release glucose into the blood by breaking down storedglycogen, and by means of gluconeogenesis. If the fall in the blood glucose level is particularly rapid or severe, other glucose sensors cause the release ofepinephrine from theadrenal glands into the blood. This has the same action as glucagon on glucose metabolism, but its effect is more pronounced.[32] In the liver glucagon and epinephrine cause thephosphorylation of the key, regulated enzymes of glycolysis,fatty acid synthesis,cholesterol synthesis, gluconeogenesis, and glycogenolysis. Insulin has the opposite effect on these enzymes.[33] The phosphorylation and dephosphorylation of these enzymes (ultimately in response to the glucose level in the blood) is the dominant manner by which these pathways are controlled in the liver, fat, and muscle cells. Thus the phosphorylation ofphosphofructokinase inhibits glycolysis, whereas its dephosphorylation through the action of insulin stimulates glycolysis.[33]
The threeregulatory enzymes arehexokinase (orglucokinase in the liver),phosphofructokinase, andpyruvate kinase. Theflux through the glycolytic pathway is adjusted in response to conditions both inside and outside the cell. The internal factors that regulate glycolysis do so primarily to provideATP in adequate quantities for the cell's needs. The external factors act primarily on theliver,fat tissue, andmuscles, which can remove large quantities of glucose from the blood after meals (thus preventinghyperglycemia by storing the excess glucose as fat or glycogen, depending on the tissue type). The liver is also capable of releasing glucose into the blood between meals, during fasting, and exercise thus preventinghypoglycemia by means ofglycogenolysis andgluconeogenesis. These latter reactions coincide with the halting of glycolysis in the liver.
In addition hexokinase andglucokinase act independently of the hormonal effects as controls at the entry points of glucose into the cells of different tissues. Hexokinase responds to theglucose-6-phosphate (G6P) level in the cell, or, in the case of glucokinase, to the blood sugar level in the blood to impart entirely intracellular controls of the glycolytic pathway in different tissues (seebelow).[33]
When glucose has been converted into G6P by hexokinase or glucokinase, it can either be converted toglucose-1-phosphate (G1P) for conversion toglycogen, or it is alternatively converted by glycolysis topyruvate, which enters themitochondrion where it is converted intoacetyl-CoA and then intocitrate. Excesscitrate is exported from the mitochondrion back into the cytosol, whereATP citrate lyase regeneratesacetyl-CoA andoxaloacetate (OAA). The acetyl-CoA is then used forfatty acid synthesis andcholesterol synthesis, two important ways of utilizing excess glucose when its concentration is high in blood. The regulated enzymes catalyzing these reactions perform these functions when they have been dephosphorylated through the action of insulin on the liver cells. Between meals, duringfasting,exercise or hypoglycemia, glucagon and epinephrine are released into the blood. This causes liver glycogen to be converted back to G6P, and then converted to glucose by the liver-specific enzymeglucose 6-phosphatase and released into the blood. Glucagon and epinephrine also stimulate gluconeogenesis, which converts non-carbohydrate substrates into G6P, which joins the G6P derived from glycogen, or substitutes for it when the liver glycogen store have been depleted. This is critical for brain function, since the brain utilizes glucose as an energy source under most conditions.[34] The simultaneously phosphorylation of, particularly,phosphofructokinase, but also, to a certain extent pyruvate kinase, prevents glycolysis occurring at the same time as gluconeogenesis and glycogenolysis.
All cells contain the enzymehexokinase, which catalyzes the conversion of glucose that has entered the cell intoglucose-6-phosphate (G6P). Since the cell membrane is impervious to G6P, hexokinase essentially acts to transport glucose into the cells from which it can then no longer escape. Hexokinase is inhibited by high levels of G6P in the cell. Thus the rate of entry of glucose into cells partially depends on how fast G6P can be disposed of by glycolysis, and byglycogen synthesis (in the cells which store glycogen, namely liver and muscles).[33][35]
Glucokinase, unlike hexokinase, is not inhibited by G6P. It occurs in liver cells, and will only phosphorylate the glucose entering the cell to form G6P, when the glucose in the blood is abundant. This being the first step in the glycolytic pathway in the liver, it therefore imparts an additional layer of control of the glycolytic pathway in this organ.[33]
Phosphofructokinase is an important control point in the glycolytic pathway, since it is one of the irreversible steps and has key allosteric effectors,AMP andfructose 2,6-bisphosphate (F2,6BP).
F2,6BP is a very potent activator of phosphofructokinase (PFK-1) that is synthesized when F6P is phosphorylated by a second phosphofructokinase (PFK2). In the liver, when blood sugar is low andglucagon elevates cAMP, PFK2 is phosphorylated byprotein kinase A. The phosphorylation inactivates PFK2, and another domain on this protein becomes active asfructose bisphosphatase-2, which converts F2,6BP back to F6P. Bothglucagon andepinephrine cause high levels of cAMP in the liver. The result of lower levels of liver F2,6BP is a decrease in activity ofphosphofructokinase and an increase in activity offructose 1,6-bisphosphatase, so that gluconeogenesis (in essence, "glycolysis in reverse") is favored. This is consistent with the role of the liver in such situations, since the response of the liver to these hormones is to release glucose to the blood.
ATP competes with AMP for the allosteric effector site on the PFK enzyme. ATP concentrations in cells are much higher than those of AMP, typically 100-fold higher,[36] but the concentration of ATP does not change more than about 10% under physiological conditions, whereas a 10% drop in ATP results in a 6-fold increase in AMP.[37] Thus, the relevance of ATP as an allosteric effector is questionable. An increase in AMP is a consequence of a decrease inenergy charge in the cell.
Citrate inhibits phosphofructokinase when testedin vitro by enhancing the inhibitory effect of ATP. However, it is doubtful that this is a meaningful effectin vivo, because citrate in the cytosol is utilized mainly for conversion toacetyl-CoA forfatty acid andcholesterol synthesis.
TIGAR, a p53 induced enzyme, is responsible for the regulation ofphosphofructokinase and acts to protect against oxidative stress.[38] TIGAR is a single enzyme with dual function that regulates F2,6BP. It can behave as a phosphatase (fructuose-2,6-bisphosphatase) which cleaves the phosphate at carbon-2 producing F6P. It can also behave as a kinase (PFK2) adding a phosphate onto carbon-2 of F6P which produces F2,6BP. In humans, the TIGAR protein is encoded byC12orf5 gene. The TIGAR enzyme will hinder the forward progression of glycolysis, by creating a build up of fructose-6-phosphate (F6P) which is isomerized into glucose-6-phosphate (G6P). The accumulation of G6P will shunt carbons into the pentose phosphate pathway.[39][40]
The final step of glycolysis is catalysed by pyruvate kinase to form pyruvate and another ATP. It is regulated by a range of different transcriptional, covalent and non-covalent regulation mechanisms, which can vary widely in different tissues.[41][42][43] For example, in the liver, pyruvate kinase is regulated based on glucose availability. During fasting (no glucose available),glucagon activatesprotein kinase A which phosphorylates pyruvate kinase to inhibit it.[44] An increase in blood sugar leads to secretion ofinsulin, which activatesprotein phosphatase 1, leading to dephosphorylation and re-activation of pyruvate kinase.[44] These controls prevent pyruvate kinase from being active at the same time as the enzymes that catalyze the reverse reaction (pyruvate carboxylase andphosphoenolpyruvate carboxykinase), preventing afutile cycle.[44] Conversely, the isoform of pyruvate kinasein found in muscle is not affected by protein kinase A (which is activated by adrenaline in that tissue), so that glycolysis remains active in muscles even during fasting.[44]
If glycolysis were to continue indefinitely, all of the NAD+ would be used up, and glycolysis would stop. To allow glycolysis to continue, organisms must be able to oxidize NADH back to NAD+. How this is performed depends on which external electron acceptor is available.
One method of doing this is to simply have the pyruvate do the oxidation; in this process, pyruvate is converted tolactate (theconjugate base of lactic acid) in a process calledlactic acid fermentation:
Pyruvate + NADH + H+ → Lactate + NAD+
This process occurs in thebacteria involved in makingyogurt (the lactic acid causes the milk to curdle). This process also occurs in animals under hypoxic (or partially anaerobic) conditions, found, for example, in overworked muscles that are starved of oxygen. In many tissues, this is a cellular last resort for energy; most animal tissue cannot tolerate anaerobic conditions for an extended period of time.
Some organisms, such as yeast, convert NADH back to NAD+ in a process calledethanol fermentation. In this process, the pyruvate is converted first to acetaldehyde and carbon dioxide, and then to ethanol.
Lactic acid fermentation and ethanol fermentation can occur in the absence of oxygen. This anaerobic fermentation allows many single-cell organisms to use glycolysis as their only energy source.
Anoxic regeneration of NAD+ is only an effective means of energy production during short, intense exercise in vertebrates, for a period ranging from 10 seconds to 2 minutes during a maximal effort in humans. (At lower exercise intensities it can sustain muscle activity indiving animals, such as seals, whales and other aquatic vertebrates, for very much longer periods of time.) Under these conditions NAD+ is replenished by NADH donating its electrons to pyruvate to form lactate. This produces 2 ATP molecules per glucose molecule, or about 5% of glucose's energy potential (38 ATP molecules in bacteria). But the speed at which ATP is produced in this manner is about 100 times that of oxidative phosphorylation. The pH in the cytoplasm quickly drops when hydrogen ions accumulate in the muscle, eventually inhibiting the enzymes involved in glycolysis.
The burning sensation in muscles during hard exercise can be attributed to the release of hydrogen ions during the shift to glucose fermentation from glucose oxidation to carbon dioxide and water, when aerobic metabolism can no longer keep pace with the energy demands of the muscles. These hydrogen ions form a part of lactic acid. The body falls back on this less efficient but faster method of producing ATP under low oxygen conditions. This is thought to have been the primary means of energy production in earlier organisms before oxygen reached high concentrations in the atmosphere between 2000 and 2500 million years ago, and thus would represent a more ancient form of energy production than the aerobic replenishment of NAD+ in cells.
The liver in mammals gets rid of this excess lactate by transforming it back into pyruvate under aerobic conditions; seeCori cycle.
Fermentation of pyruvate to lactate is sometimes also called "anaerobic glycolysis", however, glycolysis ends with the production of pyruvate regardless of the presence or absence of oxygen.
In the above two examples of fermentation, NADH is oxidized by transferring two electrons to pyruvate. However, anaerobic bacteria use a wide variety of compounds as the terminal electron acceptors incellular respiration: nitrogenous compounds, such as nitrates and nitrites; sulfur compounds, such as sulfates, sulfites, sulfur dioxide, and elemental sulfur; carbon dioxide; iron compounds; manganese compounds; cobalt compounds; and uranium compounds.
Aerobic regeneration of NAD+ and further catabolism of pyruvate
Firstly, theNADH + H+ generated by glycolysis has to be transferred to the mitochondrion to be oxidized, and thus to regenerate the NAD+ necessary for glycolysis to continue. However the inner mitochondrial membrane is impermeable to NADH and NAD+.[45] Use is therefore made of two "shuttles" to transport the electrons from NADH across the mitochondrial membrane. They are themalate-aspartate shuttle and theglycerol phosphate shuttle. In the former the electrons from NADH are transferred to cytosolicoxaloacetate to formmalate. The malate then traverses the inner mitochondrial membrane into the mitochondrial matrix, where it is reoxidized by NAD+ forming intra-mitochondrial oxaloacetate and NADH. The oxaloacetate is then re-cycled to the cytosol via its conversion to aspartate which is readily transported out of the mitochondrion. In the glycerol phosphate shuttle electrons from cytosolic NADH are transferred todihydroxyacetone to formglycerol-3-phosphate which readily traverses the outer mitochondrial membrane. Glycerol-3-phosphate is then reoxidized to dihydroxyacetone, donating its electrons toFAD instead of NAD+.[45] This reaction takes place on the inner mitochondrial membrane, allowing FADH2 to donate its electrons directly to coenzyme Q (ubiquinone) which is part of theelectron transport chain which ultimately transfers electrons to molecular oxygenO2, with the formation of water, and the release of energy eventually captured in the form ofATP.
The resulting acetyl-CoA enters thecitric acid cycle (or Krebs Cycle), where the acetyl group of the acetyl-CoA is converted into carbon dioxide by two decarboxylation reactions with the formation of yet more intra-mitochondrial NADH + H+.
The intra-mitochondrial NADH + H+ is oxidized to NAD+ by theelectron transport chain, using oxygen as the final electron acceptor to form water. The energy released during this process is used to create a hydrogen ion (or proton) gradient across theinner membrane of the mitochondrion.
Finally, the proton gradient is used to produce about 2.5ATP for every NADH + H+ oxidized in a process calledoxidative phosphorylation.[45]
Conversion of carbohydrates into fatty acids and cholesterol
The pyruvate produced by glycolysis is an important intermediary in the conversion of carbohydrates intofatty acids andcholesterol.[46] This occurs via the conversion of pyruvate intoacetyl-CoA in themitochondrion. However, this acetyl CoA needs to be transported into cytosol where the synthesis of fatty acids and cholesterol occurs. This cannot occur directly. To obtain cytosolic acetyl-CoA,citrate (produced by the condensation of acetyl CoA withoxaloacetate) is removed from thecitric acid cycle and carried across the inner mitochondrial membrane into thecytosol.[46] There it is cleaved byATP citrate lyase into acetyl-CoA and oxaloacetate. The oxaloacetate is returned to mitochondrion as malate (and then back into oxaloacetate to transfer more acetyl-CoA out of the mitochondrion). The cytosolic acetyl-CoA can be carboxylated byacetyl-CoA carboxylase intomalonyl CoA, the first committed step in thesynthesis of fatty acids, or it can be combined withacetoacetyl-CoA to form 3-hydroxy-3-methylglutaryl-CoA (HMG-CoA) which is the rate limiting step controlling thesynthesis of cholesterol.[47] Cholesterol can be used as is, as a structural component of cellular membranes, or it can be used to synthesize thesteroid hormones,bile salts, andvitamin D.[35][46][47]
Conversion of pyruvate into oxaloacetate for the citric acid cycle
Pyruvate molecules produced by glycolysis areactively transported across the innermitochondrial membrane, and into the matrix where they can either beoxidized and combined withcoenzyme A to formCO2, acetyl-CoA, and NADH,[35] or they can becarboxylated (bypyruvate carboxylase) to formoxaloacetate. This latter reaction "fills up" the amount of oxaloacetate in the citric acid cycle, and is therefore ananaplerotic reaction (from the Greek meaning to "fill up"), increasing the cycle's capacity to metabolize acetyl-CoA when the tissue's energy needs (e.g. inheart andskeletal muscle) are suddenly increased by activity.[48]In thecitric acid cycle all the intermediates (e.g. citrate, iso-citrate, alpha-ketoglutarate, succinate, fumarate, malate and oxaloacetate) are regenerated during each turn of the cycle. Adding more of any of these intermediates to the mitochondrion therefore means that that additional amount is retained within the cycle, increasing all the other intermediates as one is converted into the other. Hence the addition of oxaloacetate greatly increases the amounts of all the citric acid intermediates, thereby increasing the cycle's capacity to metabolize acetyl CoA, converting its acetate component intoCO2 and water, with the release of enough energy to form 11ATP and 1GTP molecule for each additional molecule of acetyl CoA that combines with oxaloacetate in the cycle.[48]
This article concentrates on thecatabolic role of glycolysis with regard to converting potential chemical energy to usable chemical energy during the oxidation of glucose to pyruvate. Many of the metabolites in the glycolytic pathway are also used byanabolic pathways, and, as a consequence, flux through the pathway is critical to maintain a supply of carbon skeletons for biosynthesis.[49]
The following metabolic pathways are all strongly reliant on glycolysis as a source of metabolites: and many more.
Althoughgluconeogenesis and glycolysis share many intermediates the one is not functionally a branch or tributary of the other. There are two regulatory steps in both pathways which, when active in the one pathway, are automatically inactive in the other. The two processes can therefore not be simultaneously active.[50] Indeed, if both sets of reactions were highly active at the same time the net result would be the hydrolysis of four high energy phosphate bonds (two ATP and two GTP) per reaction cycle.[50]
NAD+ is the oxidizing agent in glycolysis, as it is in most other energy yielding metabolic reactions (e.g.beta-oxidation of fatty acids, and during thecitric acid cycle). The NADH thus produced is primarily used to ultimately transfer electrons toO2 to produce water, or, whenO2 is not available, to produce compounds such aslactate orethanol (seeAnoxic regeneration of NAD+ above). NADH is rarely used for synthetic processes, the notable exception being gluconeogenesis. Duringfatty acid andcholesterol synthesis the reducing agent isNADPH. This difference exemplifies a general principle that NADPH is consumed during biosynthetic reactions, whereas NADH is generated in energy-yielding reactions.[50] The source of the NADPH is two-fold. Whenmalate is oxidatively decarboxylated by "NADP+-linked malic enzyme"pyruvate,CO2 and NADPH are formed. NADPH is also formed by thepentose phosphate pathway which converts glucose into ribose, which can be used in synthesis ofnucleotides andnucleic acids, or it can be catabolized to pyruvate.[50]
Cellular uptake of glucose occurs in response to insulin signals, and glucose is subsequently broken down through glycolysis, lowering blood sugar levels. However, insulin resistance or low insulin levels seen in diabetes result in hyperglycemia, where glucose levels in the blood rise and glucose is not properly taken up by cells. Hepatocytes further contribute to this hyperglycemia throughgluconeogenesis. Glycolysis in hepatocytes controls hepatic glucose production, and when glucose is overproduced by the liver without having a means of being broken down by the body, hyperglycemia results.[51]
Glycolytic mutations are generally rare due to importance of the metabolic pathway; the majority of occurring mutations result in an inability of the cell to respire, and therefore cause the death of the cell at an early stage. However, some mutations (glycogen storage diseases and otherinborn errors of carbohydrate metabolism) are seen with one notable example beingpyruvate kinase deficiency, leading to chronic hemolytic anemia.[citation needed]
Malignant tumor cells perform glycolysis at a rate that is ten times faster than their noncancerous tissue counterparts.[53] During their genesis, limited capillary support often results in hypoxia (decreased O2 supply) within the tumor cells. Thus, these cells rely on anaerobic metabolic processes such as glycolysis for ATP (adenosine triphosphate). Some tumor cells overexpress specific glycolytic enzymes which result in higher rates of glycolysis.[54] Often these enzymes are Isoenzymes, of traditional glycolysis enzymes, that vary in their susceptibility to traditional feedback inhibition. The increase in glycolytic activity ultimately counteracts the effects of hypoxia by generating sufficient ATP from this anaerobic pathway.[55] This phenomenon was first described in 1930 byOtto Warburg and is referred to as theWarburg effect. TheWarburg hypothesis claims that cancer is primarily caused by dysfunctionality in mitochondrial metabolism, rather than because of the uncontrolled growth of cells.A number of theories have been advanced to explain the Warburg effect. One such theory suggests that the increased glycolysis is a normal protective process of the body and that malignant change could be primarily caused by energy metabolism.[56]
There is ongoing research to affect mitochondrial metabolism and treat cancer by reducing glycolysis and thus starving cancerous cells in various new ways, including aketogenic diet.[59][60][61]
The diagram below shows human protein names. Names in other organisms may be different and the number ofisozymes (such as HK1, HK2, ...) is likely to be different too.
Click on genes, proteins and metabolites below to link to respective articles.[§ 1]
Some of the metabolites in glycolysis have alternative names and nomenclature. In part, this is because some of them are common to other pathways, such as theCalvin cycle.
The intermediates of glycolysis depicted in Fischer projections show the chemical changing step by step. Such image can be compared to polygonal model representation.[62]
Glycolysis - Structure of anaerobic glycolysis components showed using Fischer projections, left, and polygonal model, right. The compounds correspond to glucose (GLU), glucose 6-phosphate (G6P), fructose 6-phosphate (F6P), fructose 1,6-bisphosphate ( F16BP), dihydroxyacetone phosphate (DHAP), glyceraldehyde 3-phosphate(GA3P), 1,3-bisphosphoglycerate (13BPG), 3-phosphoglycerate (3PG), 2-phosphoglycerate (2PG), phosphoenolpyruvate (PEP), pyruvate (PIR), and lactate (LAC). The enzymes which participate of this pathway are indicated by underlined numbers, and correspond to hexokinase (1), glucose-6-phosphate isomerase (2), phosphofructokinase-1 (3), fructose-bisphosphate aldolase (4), triosephosphate isomerase (5), glyceraldehyde-3-phosphate dehydrogenase (5), phosphoglycerate kinase (7), phosphoglycerate mutase (8), phosphopyruvate hydratase (enolase) (9), pyruvate kinase (10), and lactate dehydrogenase (11). The participant coenzymes (NAD+, NADH + H+, ATP and ADP), inorganic phosphate,H2O andCO2 were omitted in these representations. The phosphorylation reactions from ATP, as well the ADP phosphorylation reactions in later steps of glycolysis are shown as ~P respectively entering or going out the pathway. The oxireduction reactions using NAD+ or NADH are observed as hydrogens "2H" going out or entering the pathway.
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