Glycolysis Flashcards
Describe aerobic glycolysis (overall reaction purpose, prep/payoff phases + enzymes, fates of pyruvate)
1 mole glucose -> 2 mole of pyruvate
Degradation of glucose into pyruvate, requires 10 enzymes
Goal: unlock potential energy in glucose by making high energy intermediates that can drive substrate level phosphorylation.
Pyruvate fates: aerobic metabolism, anaerobic respiration and ethanol fermentation.
Preparatory phase: expense of ATP to phosphorylate and degrade glucose to trioses
-Activation of carbon through 2 successive phosphorylations (expends ATP)
-Cleaving of ring to generate 2x 3 carbons molecules called trioses.
Enzymes: acronyms are okay
-Hexokinase/Glucokinase (HK/GK)
-Phosphoglucose isomerase (PGI)
-Phosphofructokinase-1 (PFK1)
-Aldolase (FBA)
-Triose phosphate isomerase (TPI)
Payoff phase: trioses are oxidized and dephosphorylated to produce ATP by substrate level phosphorylation
-Oxidation of prepares trioses
-Substrate level phosphorylation of ADP by using phosphoryl groups. (Generates 4 ATP and 2 pyruvate)
Enzymes:
-Glyceraldehyde-3-phosphate dehydrogenase (G3PDH)
-phosphoglycerate kinase (PGK)
-phosphoglycerate mutase (PGM)
-enolase (ENO)
-pyruvate kinase (PK)
Describe how glucose enters a cell and the different fates of glucose once it is uptaken by a cell.
Describe particularities of GLUT and the concentration gradient. How the concentration gradient maintained inside the cell in the fed state?
Difference between hexokinase and glucokinase?
Why is Mg2+ present for the 1st reaction of glycolysis?
Glucose enters cell cytosol from extracellular milieu through GLUT (follows concentration gradient).
Fates of glucose: G6P (metabolic intersection)
1. G6P -> NADPH or Ribose-5-Phosphate in PPP (genesis of ribose units)
2. Glycogenesis (energy storage)
3. Energy metabolism through production of other metabolites, cellular respiration, etc.. (production of energy)
4. Glycoproteins (structure)
GLUT: uniporter of glucose (transports one direction) depending on glucose concentration gradient
-Fed state: glucose enters cell
-Fasting state: glucose exits cell
Concentration gradient maintained with first step of glycolysis
Glucose -> Glucose-6-phosphate by hexokinase/glucokinase
-Maintains gradient to keep GLUT working
-starts glycolysis.
-G6P is a metabolic intersection, can lead to different fates
-G6P is negatively charged with Pi, so it has electrostatic repulsion from negative PL, and stays trapped in the cell.
They both catalyze the same reaction (G->G6P), but
Hexokinase: found in most tissue
Glucokinase: unique to the liver
Mg2+ present as cofactor in all ATP reactions.
-binds to Pi of ATP to make them more available for transfer to glucose.
Why is glucose an important source of energy? Compared to palmitate?
What is the formula of cellular respiration?
Why is stockpiling glucose advantageous?
Glucose is rich in potential energy (stored in covalent bonds), and is thus a good fuel. Its complete oxidation to CO2 and H20 yields -2840kJ/mol of energy.
Palmitate is richer in potential energy, but glucose is the key energy source in the body, especially for brain tissue.
Glucose + 6O2 -> 6CO2 + 6H2O + 36 ATP
Polymerization and storage of glucose as glycogen is advantageous. Monomeric hexose units are mobilized from glycogen to make ATP and other molecules in times of increased energy demand.
What do these different enzymes do?
Kinase, isomerase, aldolase, dehydrogenase, mutase, enolase
If a reaction is irreversible, what does that mean about its Gibb’s free energy?
Kinase: phosphorylates
Isomerase: changes structure
Aldolase: cleaves carbon chain
Dehydrogenase: redox reaction (moving electrons)
Mutase: moves functional group within a molecule
Enolase: involved in hydrations
Irreversible reaction: gibbs free energy is too high for the reaction to reverse.
What is the difference between aerobic and anaerobic glycolysis?
What are allosteric regulators? What type of reactions do they usually regulate?
Aerobic: occurs in presence of mitochondria, pyruvate burned to drive oxidative phosphorylation.
Anaerobic: occurs in erythrocytes and muscle cells, pyruvate burned to drive substrate level phosphorylation by producing lactate as a byproduct
Pyruvate + NADH -> Lactate + NAD+ by LACTATE DEHYDROGENASE
(type II muscle cells do this too during intense exercise to rapidly generate ATP)
Allosteric regulators: molecules that bind to an enzyme at a site other than the active site, known as the allosteric site. This binding induces a conformational change in the enzyme that can either enhance (activate) or inhibit (deactivate) its activity.
Typically regulate irreversible reactions, involving ATP. These are the reactions that produce intermediates of metabolic intersections (G6P) ** is this true?
Describe the preparatory phase of glycolysis
-substrates and products
-enzymes
-allosteric regulators of enzymes
-investment and splitting phase
-irreversible reactions
-prep products
Preparatory phase of glycolysis
Investment phase:
1. Glucose + ATP + Mg2+ -> Glucose-6-Phosphate (G6P) +ADP
-irreversible
-hexokinase/glucokinase (HK/GK)
allosteric regulators:
for GK: +glucose, +insulin, -glucagon, -fructose-6-phosphate
for HK: -glucose-6-phosphate
- G6P -> Fructose-6-Phosphate (F6P)
-phosphoglucose isomerase (PGI)
allosteric regulators: none - F6P + ATP + Mg2+ -> Fructose-1,6-Biphosphate (F16BP) + ADP
-irreversible
-phosphofructokinase (PFK1)
allosteric regulators: +AMP/ADP, +F26BP***, -ATP, -citrate (both from CAC)
Splitting phase:
4. F16BP -> Dihydroxyacetone (DHAP) + Glyceraldehyde-3-Phosphate (G3P)
-aldolase (FBA)
allosteric regulators: none
- DHAP -> G3P
-triose phosphate isomerase (TPI)
allosteric regulators: none
Overall product:
-2 ATP
Describe the payoff phase of glycolysis
-substrates and products
-enzymes
-allosteric regulators of enzymes
-recoup phase
-irreversible reactions
-payoff products
-net product of glycolysis
What is the 11th step of glycolysis?
Payoff phase of glycolysis (happens 2x)
Recoup phase:
6. G3P +NAD+ +Pi -> 1,3-Biphosphoglycerate (13BPG) + NADH
-Glyceraldehyde-3-P dehydrogenase (G3PDH)
allosteric regulators: none
- 13BPG + ADP -> 3-phosphoglycerate (3PG) + ATP
-Phospholycertae kinase (PGK)
-reversible reaction (Exception)
allosteric regulators: none - 3PG -> 2-phosphoglycerate (2PG)
-Phosphoglycerate mutase (PGM)
allosteric regulators: none - 2PG -> Phosphoenolpyruvate (PEP)
-enolase
allosteric regulators: none - PEP + ADP -> Pyruvate + ATP
-Pyruvate kinase (PK)
-irreversible
allosteric regulators: +insulin, +F16BP, -alanine, -ATP, -glucagon
Overall products:
4 ATP, 2 NADH, 2 Pyruvate
Net products of all glycolysis:
2 ATP, 2 NADH, 2 Pyruvate
- Pyruvate + NADH -> Lactate + NAD+
-lactate dehydrogenase (LDH)
Regeneration of NAD+ to go back up the chain
Name the irreversible reactions of glycolysis
What are the high energy intermediates?
- Glucose -> Glucose-6-phosphate
- Fructose-6-phosphate -> Fructose-1,6-biphosphate
- Phosphoenolpyruvate -> pyruvate
High energy intermediates: molecules containing high potential energy.
G6P, F16BP, 13BPG, PEP
How is glucose metabolism modified in cancer cells?
What is the Warbug effect?
Cancer cells are glucose loving tissues, that will rewire metabolic pathways by…
Overactivating glycolysis to produce ATP extremely rapidly, which significantly increases lactate production.
MCT transporter in cell membrane exports lactate from the cell to increase tumor biomass and cell proliferation.
-Glycolysis outpaces mitochondria
-Pyruvate doesn’t enter mitochondria
Warbug effect: cancer cells preferentially utilizing glucose for energy even in the presence of oxygen. This leads to increased glucose levels
How can cancer be clinically detected using glucose metabolism?
How can brown fat tissue be detected?
Fluorodeoxyglucose (FDG) is a glucose analog, with a missing OH in its C2 position and a fluoride group (positron emitter).
FDG will be uptaken by cells, and converted to FDG-6P by HK. The OH on its C2 will prevent further metabolism, causing it to accumulate in gluose-loving cells.
Ex: cancer cells, brain, bladder, heart, kidney, etc…
High glucose demands of cancer cells allow its diagnosis:
Positron emission tomography (PET scans) can detect positron emission from accumulated FDG, allowing detection of cancer tumours.
By making patients cold and injecting them with FDG, accumulation of FDG in activated brown fat tissue will allow detection with PET scans.
AMP is an important allosteric regulator of metabolism. Explain why.
For which enzymes in AMP an allosteric activator?
For which enzymes is ATP an allosteric inhibitor?
What is another way to modulate metabolic pathways?
ATP is not a good allosteric regulator, because its concentration does not vary much in the cell.
ATP -> ADP + Pi
ADP + ADP -> AMP + ATP
AMP is the cell’s master fuel gauge.
High AMP = high energy demands, promotes ATP production.
Low AMP = low energy demandes, inhibits ATP production.
AMP is an allosteric activator for PFK1
ATP is an allosteric inhibitor for PFK1 and pyruvate kinase
Gene transcription activation and inhibition is another way metabolic pathways can be modulated.
Describe transcriptional regulation of glycolysis through ChREBP
What gene transcription response is expected from glycolytic gene activation? Why?
Would glycolytic gene transcription be overactivated in cancer cells? why?
ChREBP: Carbohydrate Response Element Binding Protein
Sits in cytoplasm until change in nutritional status of the cell occurs (glucose enters and is metabolized).
- G -> G6P
- G6P -> F6P (glycolysis)
- F6P -> F26BP/X5P (PPP)
These three metabolites bind to ChREBP and trigger nuclear localization.
-X5P (xylulose-5-phosphate) from PPP dephosphorylates ChREBP.
-G6P allosterically activates ChREBP
- ChREBP enters nucleus, binds to ChoRE gene, with OGT and CBP/p300 proteins
- Activates gene transcription for genes involved in glycolytic response and lipogenesis response.
As glucose enters cells, it will be converted to fat for storage under insulin’s effects, which is why lipogenesis genes are also activated.
In a cancer cells, this process would be overactivated because
-High rates of glycolysis increase metabolites activating this gene transcription pathway
-Lipogenesis is crucial for cell membrane synthesis and cell proliferation.
What is the hypoxic response?
What is hypoxia and normoxia?
What is the cell’s response in hypoxia and normoxia?
What happens to mitochondria in response to normoxia and hypoxia?
Is HIF-1a oncogenic?
Hypoxic response: cell’s ability to respond to oxygen availability.
Hypoxia: oxygen defiency
Normoxia: normal oxygen levels
Cell response:
Normoxia: normal circumstances
1. PHD utilizes O2, Fe2+ and Ascorbate to hydroxylate proline on HIF-1a
2. hydroxylated HIF-1a then goes to 26S proteosome for proteasomal degradation.
Hypoxia: low oxygen levels
1. PHD doesn’t have O2 to utilize to hydroxylate HIF-1a, so it stabilizes.
2.HIF-1a combines with p300/CBP and HIF-1B nuclear proteins and activates HIF gene in the nucleus
3. Activation of gene transcription for glycolysis (ATP production), vascularization, erythropeosis and Fe uptake, all in the goal of increasing oxygen rates.
Mitochondria:
Normoxia:
-PGC-1a and PCG-1b genes transcribed for mitochondrial biogenesis.
Hypoxia:
-HIF-1a degrades PGC-1a and PCG-1b genes, resulting in less mitochondrial biogenesis
HIF-1a has oncogenic tendencies, and can promote the development of cancer, through tumor vascularization and cancer cell glycolysis. It promotes various cellular processes that support tumor growth and survival, especially in low-oxygen (hypoxic) environments
