Glycolysis stands as a cornerstone of cellular metabolism, a fundamental pathway employed by every living cell to extract energy from glucose. This intricate process not only fuels cellular activities but also generates crucial intermediate compounds vital for a myriad of other metabolic routes. Beyond glucose, glycolysis adeptly handles other simple sugars like fructose and galactose, seamlessly integrating them into the energy-generating machinery of the cell [1].
Glycolysis: A Cytoplasmic Affair
The pivotal question, Where Does Glycolysis Take Place In The Cell?, has a straightforward answer: glycolysis unfolds within the cytoplasm. This gel-like substance filling the cell is the stage for the entire ten-step biochemical ballet that transforms a single six-carbon glucose molecule into two three-carbon pyruvate molecules. The subsequent journey of pyruvate, however, is contingent on the cellular environment, specifically the presence of mitochondria and oxygen.
In cells equipped with mitochondria, often dubbed the powerhouses of the cell, and under oxygen-rich (aerobic) conditions, pyruvate embarks on the next phase of energy extraction. It is transported into the mitochondria where the pyruvate dehydrogenase complex orchestrates its conversion into Acetyl-CoA. This molecule then enters the Tricarboxylic Acid (TCA) cycle, also known as the Krebs cycle, setting the stage for the electron transport chain, the primary driver of ATP (adenosine triphosphate) production, the cell’s energy currency.
However, in the absence of oxygen (anaerobic conditions) or in cells lacking mitochondria altogether, such as red blood cells (erythrocytes), an alternative pathway, anaerobic glycolysis, takes precedence. Here, pyruvate is reduced to lactate by lactate dehydrogenase, concurrently reoxidizing NADH back to NAD+. This seemingly simple step is crucial because it regenerates NAD+, essential for the continuation of glycolysis itself, allowing for a continuous, albeit less efficient, ATP production in the absence of oxygen. During aerobic glycolysis, the NADH generated is not wasted; instead, it is shuttled into the mitochondria via systems like the malate-aspartate shuttle or glycerol phosphate shuttle. Within the mitochondria, this NADH contributes its electrons to the electron transport chain, maximizing ATP generation [1], [2].
A Step-by-Step Journey Through Glycolysis in the Cytoplasm
Aerobic glycolysis, despite its name suggesting oxygen dependence, does not directly require oxygen in its ten cytoplasmic steps. Oxygen’s role becomes crucial later in reoxidizing NADH in the mitochondria. Let’s dissect each step of this fascinating pathway occurring in the cell’s cytoplasm [1]:
Step 1: Glucose Phosphorylation – Trapping Glucose in the Cell
Upon entering the cell, glucose encounters hexokinase, the first gatekeeper of glycolysis. This enzyme swiftly adds a phosphate group to glucose, transforming it into glucose-6-phosphate. This phosphorylation is an irreversible reaction, consuming one molecule of ATP and, importantly, trapping glucose within the cellular confines. Hexokinase exhibits broad specificity, welcoming various six-carbon sugars. However, liver and pancreatic beta cells host a specialized isozyme, glucokinase, which is more glucose-specific.
Step 2: Isomerization – Preparing for the Split
Glucose-6-phosphate, an aldose sugar, undergoes isomerization into fructose-6-phosphate, a ketose sugar, courtesy of phosphoglucose isomerase. This reaction is readily reversible, setting the stage for the next regulatory checkpoint.
Step 3: The Second Phosphorylation – A Committed Step
Fructose-6-phosphate is further phosphorylated by phosphofructokinase-1 (PFK-1) to yield fructose-1,6-bisphosphate. This step is not only irreversible but also the rate-limiting and primary regulatory point of glycolysis. It is the second ATP-consuming step, solidifying the commitment of glucose towards glycolysis.
Step 4: Sugar Splitting – Aldolase Action
Fructose-1,6-bisphosphate is cleaved by aldolase into two three-carbon molecules: dihydroxyacetone phosphate (DHAP) and glyceraldehyde-3-phosphate (G3P). This reversible and unregulated reaction is crucial as it sets up the pathway for processing glucose as two three-carbon units. The liver isoform, aldolase B, also plays a role in fructose metabolism by cleaving fructose-1-phosphate.
Step 5: Isomer Interconversion – Balancing the Pathway
Triose phosphate isomerase catalyzes the reversible interconversion between DHAP and glyceraldehyde-3-phosphate. This ensures that DHAP is efficiently converted to G3P, meaning that from one glucose molecule, two molecules of glyceraldehyde-3-phosphate are available to proceed through the subsequent steps of glycolysis.
Step 6: Oxidation and Phosphorylation – Generating NADH and High-Energy Intermediate
Glyceraldehyde-3-phosphate dehydrogenase orchestrates the oxidation of glyceraldehyde-3-phosphate, leading to the formation of 1,3-bisphosphoglycerate. This is the first redox reaction in glycolysis. Here, NAD+ is reduced to NADH, while glyceraldehyde-3-phosphate’s aldehyde group is oxidized to a carboxyl group and simultaneously phosphorylated. The availability of NAD+ is crucial; its regeneration is accomplished in the mitochondria under aerobic conditions and by lactate dehydrogenase under anaerobic conditions.
Step 7: ATP Generation – The First Energy Payoff
1,3-bisphosphoglycerate is converted to 3-phosphoglycerate by phosphoglycerate kinase. This step marks the first substrate-level phosphorylation in glycolysis, where the high-energy phosphate group from 1,3-BPG is transferred to ADP, generating ATP. This step yields two ATP molecules per glucose molecule, as we are now processing two three-carbon units. Interestingly, a portion of 1,3-BPG can be diverted to form 2,3-bisphosphoglycerate (2,3-BPG) by bisphosphoglycerate mutase, a molecule crucial for regulating oxygen delivery, especially under hypoxic conditions.
Step 8: Phosphate Group Shift – Preparing for Dehydration
Phosphoglycerate mutase catalyzes the reversible isomerization of 3-phosphoglycerate to 2-phosphoglycerate, shifting the phosphate group from carbon 3 to carbon 2. This repositioning is essential for the subsequent step.
Step 9: Dehydration – Creating a High-Energy Enol Phosphate
2-phosphoglycerate is dehydrated by enolase to form phosphoenolpyruvate (PEP). This seemingly simple dehydration step creates a high-energy enol phosphate bond, poised for ATP generation in the final step.
Step 10: Pyruvate Formation and ATP Generation – The Final Payoff
Pyruvate kinase catalyzes the irreversible conversion of phosphoenolpyruvate to pyruvate, the end product of glycolysis. This final substrate-level phosphorylation step generates another two ATP molecules per glucose molecule.
In summary, glycolysis, occurring entirely within the cytoplasm, converts one glucose molecule into two pyruvate molecules, yielding a net gain of two ATP molecules and two NADH molecules, irrespective of whether pyruvate subsequently enters the aerobic or anaerobic pathway.
Regulatory Mechanisms of Glycolysis
Glycolysis is not a runaway train; it’s meticulously controlled to meet cellular energy demands. Regulation occurs at multiple levels: covalent modification of key enzymes, allosteric modulation, and hormonal control.
A key regulatory enzyme is the bifunctional enzyme PFK2/Fructose bisphosphatase. Its kinase activity synthesizes fructose-2,6-bisphosphate (F2,6-BP), a potent allosteric activator of PFK-1, the rate-limiting enzyme of glycolysis. The balance between insulin and glucagon dictates F2,6-BP levels. High insulin levels, indicative of glucose abundance, activate protein phosphatase, dephosphorylating PFK2, thus activating its kinase domain. This surge in F2,6-BP boosts glycolysis.
Conversely, high glucagon levels, signaling glucose scarcity, trigger a cascade involving cAMP and protein kinase A. This kinase phosphorylates the bifunctional enzyme, inhibiting PFK2 activity and activating its phosphatase domain. Consequently, F2,6-BP levels plummet, glycolysis is suppressed, and gluconeogenesis (glucose synthesis) is favored.
Hormones, particularly insulin and glucagon, exert significant control. Insulin, released in response to elevated blood glucose, upregulates glucokinase, PFK-1, and pyruvate kinase – the three irreversible enzymes of glycolysis – ensuring efficient glucose utilization. Simultaneously, low glucagon levels prevent counterproductive gluconeogenesis. Long-term hormonal control via gene transcription is critical in states like fasting, starvation, and diabetes, where the insulin/glucagon ratio is chronically altered. In these scenarios, the synthesis of glucokinase, PFK-1, and pyruvate kinase is diminished at the gene level [1], [3], [1].
Clinical Relevance of Glycolysis
Dysregulation of glycolysis is implicated in various diseases, highlighting its critical role in human health.
Glucokinase Deficiency: Both glucokinase and hexokinase phosphorylate glucose, but glucokinase, primarily in the liver and pancreatic beta cells, has a lower glucose affinity and operates mainly at high glucose concentrations. In beta cells, glucokinase acts as a glucose sensor, modulating insulin secretion. Mutations in glucokinase can lead to Maturity-Onset Diabetes of the Young type 2 (MODY2), a form of diabetes characterized by mild hyperglycemia [4], [5]. Severe glucokinase deficiency can result in neonatal diabetes mellitus [6], [7], [8].
2,3-Bisphosphoglycerate (2,3-BPG): Red blood cells utilize glycolysis for energy and produce 2,3-BPG. Elevated 2,3-BPG levels occur in response to low oxygen availability (high altitude, lung diseases). 2,3-BPG reduces hemoglobin’s oxygen affinity, facilitating oxygen release to tissues. This is reflected in a rightward shift of the oxygen dissociation curve [9].
Pyruvate Kinase Deficiency: This autosomal recessive disorder, stemming from PKLR gene mutations, impairs pyruvate kinase activity, particularly affecting red blood cells, which rely solely on glycolysis for ATP. ATP depletion in RBCs compromises membrane integrity, leading to hemolysis, causing fatigue, shortness of breath, and jaundice due to bilirubin accumulation. Damaged RBCs become echinocytes (spiky cells). The body compensates by increasing 2,3-BPG to improve oxygen delivery, but this doesn’t resolve the ATP deficit [10]. Liver pyruvate kinase deficiency is less severe due to mitochondrial ATP production.
Pyruvate Kinase and Cancer: Pyruvate kinase is often upregulated in cancer cells and rapidly proliferating embryonic cells. Cancer cells exhibit the Warburg effect, favoring aerobic glycolysis even with sufficient oxygen. This metabolic shift, diverting pyruvate to lactate production, may provide cancer cells with a rapid energy source, biosynthetic precursors, and an acidic microenvironment conducive to invasion and metastasis. The M2 isoform of pyruvate kinase is particularly implicated in cancer [11], [12], [13], [14], [15], [16].
Review Questions
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References
1.Dashty M. A quick look at biochemistry: carbohydrate metabolism. Clin Biochem. 2013 Oct;46(15):1339-52. [PubMed: 23680095]
2.Niu X, Arthur P, Abas L, Whisson M, Guppy M. Carbohydrate metabolism in human platelets in a low glucose medium under aerobic conditions. Biochim Biophys Acta. 1996 Oct 24;1291(2):97-106. [PubMed: 8898869]
3.Rui L. Energy metabolism in the liver. Compr Physiol. 2014 Jan;4(1):177-97. [PMC free article: PMC4050641] [PubMed: 24692138]
4.Tattersall RB. Mild familial diabetes with dominant inheritance. Q J Med. 1974 Apr;43(170):339-57. [PubMed: 4212169]
5.Osbak KK, Colclough K, Saint-Martin C, Beer NL, Bellanné-Chantelot C, Ellard S, Gloyn AL. Update on mutations in glucokinase (GCK), which cause maturity-onset diabetes of the young, permanent neonatal diabetes, and hyperinsulinemic hypoglycemia. Hum Mutat. 2009 Nov;30(11):1512-26. [PubMed: 19790256]
6.Froguel P, Vaxillaire M, Sun F, Velho G, Zouali H, Butel MO, Lesage S, Vionnet N, Clément K, Fougerousse F. Close linkage of glucokinase locus on chromosome 7p to early-onset non-insulin-dependent diabetes mellitus. Nature. 1992 Mar 12;356(6365):162-4. [PubMed: 1545870]
7.Hattersley AT, Turner RC, Permutt MA, Patel P, Tanizawa Y, Chiu KC, O’Rahilly S, Watkins PJ, Wainscoat JS. Linkage of type 2 diabetes to the glucokinase gene. Lancet. 1992 May 30;339(8805):1307-10. [PubMed: 1349989]
8.Froguel P, Zouali H, Vionnet N, Velho G, Vaxillaire M, Sun F, Lesage S, Stoffel M, Takeda J, Passa P. Familial hyperglycemia due to mutations in glucokinase. Definition of a subtype of diabetes mellitus. N Engl J Med. 1993 Mar 11;328(10):697-702. [PubMed: 8433729]
9.Benesch R, Benesch RE. The effect of organic phosphates from the human erythrocyte on the allosteric properties of hemoglobin. Biochem Biophys Res Commun. 1967 Jan 23;26(2):162-7. [PubMed: 6030262]
10.Grace RF, Zanella A, Neufeld EJ, Morton DH, Eber S, Yaish H, Glader B. Erythrocyte pyruvate kinase deficiency: 2015 status report. Am J Hematol. 2015 Sep;90(9):825-30. [PMC free article: PMC5053227] [PubMed: 26087744]
11.WARBURG O. On the origin of cancer cells. Science. 1956 Feb 24;123(3191):309-14. [PubMed: 13298683]
12.Mazurek S, Boschek CB, Hugo F, Eigenbrodt E. Pyruvate kinase type M2 and its role in tumor growth and spreading. Semin Cancer Biol. 2005 Aug;15(4):300-8. [PubMed: 15908230]
13.Christofk HR, Vander Heiden MG, Wu N, Asara JM, Cantley LC. Pyruvate kinase M2 is a phosphotyrosine-binding protein. Nature. 2008 Mar 13;452(7184):181-6. [PubMed: 18337815]
14.Gupta V, Wellen KE, Mazurek S, Bamezai RN. Pyruvate kinase M2: regulatory circuits and potential for therapeutic intervention. Curr Pharm Des. 2014;20(15):2595-606. [PubMed: 23859618]
15.Olson KA, Schell JC, Rutter J. Pyruvate and Metabolic Flexibility: Illuminating a Path Toward Selective Cancer Therapies. Trends Biochem Sci. 2016 Mar;41(3):219-230. [PMC free article: PMC4783264] [PubMed: 26873641]
16.Gatenby RA, Gillies RJ. Why do cancers have high aerobic glycolysis? Nat Rev Cancer. 2004 Nov;4(11):891-9. [PubMed: 15516961]
Disclosure: Jeffrey Naifeh declares no relevant financial relationships with ineligible companies.
Disclosure: Manjari Dimri declares no relevant financial relationships with ineligible companies.
Disclosure: Matthew Varacallo declares no relevant financial relationships with ineligible companies.
