Glycolysis

• What does hexokinase do?

  Phosphorylates glucose into glucose 6-phosphate using ATP
• What does phosphoglucose isomerase do?


  Turns the six-membered ring of glucose 6-phosphate into the five-membered ring of fructose 6-phosphate
• What does phosphofructokinase do?

  Phosphorylates fructose 6-phosphate into fructose 1,6-biphosphate using ATP.
• What does aldolase do?

  Cuts fructose 1,6-biphosphate in half into glyceraldehyde 3-phosphate and dihydroxyacetone phosphate
• What does triose phosphate isomerase do?

  Catalyzes the reversible isomerization / conversion of dihydroxyacetone to glyceraldehyde 3-phoshate
• How does hexokinase reduce the hydrolysis of ATP?

  The induced fit of the enzyme brings the two lobes of the enzyme closer together, displacing water from the active site and preventing the loss of ATP being used in hydrolysis.
• Which enzymes catalyze reversible reactions in the first stage of glycolysis (trapping and preparing)?

  1. phosphoglucose isomerase
  2. triose phosphate isomerase
• Once glucose is phosphorylated (glucose 6-phosphate), it cannot leave the cell
• What does glyceraldehyde 3-phosphate dehydrogenase do and use?

  Catalyzes glyceraldehyde 3-phosphate into 1,3-bisphosphoglycerate by oxidizing the aldehyde and phosphate while reducing NAD+
• What does phosphoglycerate kinase do?

  Catalyzes 1,3-bisphosphoglycerate into 3-phosphoglycerate by removing a phosphate group from 1,3-BPG and adding it to ADP. ATP is gained in this reaction.
• Sodium-glucose linked transporter: symporter that carries sodium and glucose / galactose into the cell
• How many pyruvate are made from 1 molecule of glucose?

  2
• What is the goal of the first stage of glycolysis?

  To trap glucose in the cell by phosphorylation and transforming it into molecules that can be better broken down
• Why might glucose would have become such a prominent fuel source as opposed to other monosaccharide?

  Glucose is the most stable hexose because the hydroxyl groups and the hydroxymethyl group are all in the equatorial position minimizing steric hinderances. Also, glucose has a low tendency to glycosylate proteins as opposed to other monosaccharides in their open-chain form where their carbonyl groups can covalently modify the amino groups of proteins, often making the proteins function less efficiently.
• 1. Goal of the first stage of glycolysis is to trap the glucose in the cell and form a compound that can be readily cleaved into phosphorylated 3- carbon units. This stage is complete when 1,6-bisphosphate is cleaved into 2 phosphorylated three-carbon molecules (i.e., everything in 2nd stage of glycolysis, and glyceraldehyde-3-phosphate and dihydroxyacetone phosphate). No ATP is made in stage 1 of glycolysis.
  2. In the second stage of glycolysis, ATP is harvested when the phosphorylated three-carbon units are oxidized to pyruvate
• Which enzyme traps glucose in the cell and begins glycolysis?

  Hexokinase. Hexokinase phosphorylates glucose by ATP to form glucose-6-phosphate. This step is notable for two reasons:

  1. glucose-6-phosphate cannot pass through the membrane to the extracellular side because it is not a substrate for the glucose transporters
  2. the addition of the phosphoryl group facilitates the metabolism of glucose to phosphorylated three-carbon fragments with high phosphoryl-transfer potential
• Kinases are enzymes that catalyze the transfer of a phosphoryl group from ATP to an acceptor. Kinases require Mg2+ for activity (or another divalent metal ion such as Mn2+) to form a complex with ATP.
• Describe the conformational changes of hexokinase when it binds to glucose?

  Induced fit; the two lobes of hexokinase move towards each other when glucose is bound, and when the cleft between the lobes closes, the bound glucose becomes surrounded by protein except for the carbon atom that will accept the phosphoryl group from ATP.

  • Water is removed from the active site which enhances the specificity of hexokinase by:
  • preventing side reactions of water with other molecules in the active site
  • preventing water from competing with glucose for binding to the active site
• What does phosphoglucose isomerase do?

  Catalyzes the isomerization of glucose 6-phosphate to fructose 6-phosphate by converting the aldose of glucose into an ketose.
• At equilibrium, 96% of the triose phosphate is dihydroxyacetone phosphate (DHAP). However once DHAP is formed, it is readily converted to glyceraldehyde 3-phosphate by triose phosphate isomerase. This conversion effectively removes DHAP from the system, driving the equilibrium of the reversible reaction towards the formation of more DHAP.
• In the first stage of glycolysis how much ATP has been spent?

  2
• What does glyceraldehyde 3-phosphate dehydrogenase do?

  Catalyzes the conversion of glyceraldehyde 3-phosphate into 1,3-bisphosphoglycerate (1,3-BPG) in an oxidation-reduction reaction. NAD+ is reduced to NADH in this reaction.
• Dehydrogenases are enzymes that catalyze oxidation-reduction reactions, often transferring a hydride ion from a donor molecule to NAD+ or transferring a hydride ion from NADH to an acceptor molecule
• which molecules are higher energy molecules than ATP in glycolysis?

  1,3-BPG, PEP, and creatine phosphate
• How is 1,3-BPG converted to 3-phosphoglycerate if it is thermodynamically unfavourable?

  A high energy thioester intermediate linked to glyceraldehyde 3-phosphate dehydrogenase lowers the activation energy of the formation of the 3-phosphoglycerate (acyl-phosphate) product. The intermediate undergoes covalent catalysis
• How is ATP formed from phosphoryl transfer from 1,3-bisphosphoglycerate?

  1,3-BPG is an energy-rich molecule with a greater phosphoryl-transfer potential than ATP, so 1,3-BPG can be used to power the synthesis of ATP from ADP and Pi. Phosphoglycerate kinase catalyzes the transfer of phosphoryl to ADP from 1,3-BPG. 2 ATPs formed because two 1,3-BPG were made
• What does phosphoglycerate mutase do?

  Catalyzes conversion of 3-phosphoglycerate to 2-phosphoglycerate by shifting the position of the phosphoryl group.
• Mustases catalyze the shift of a functional group from one position to another within the same molecule
• Which reaction has a dehydration reaction and why does it matter?

  Conversion of 2-phosphoglycerate to phosphoenolpyruvate by enolase is the dehydration reaction. The dehydration elevates the phosphoryl transfer potential since enol phosphates are highly unstable and will convert to a more stable ketone form (pyruvate) after the phosphoryl group is donated to ADP.
• How does phosphoenolpyruvate conversion generate ATP?

  The highly unstable enol form gives the molecule a high phosphoryl transfer potential, thus it likely to donate its phosphoryl group to ADP to generate ATP using the enzyme pyruvate kinase to catalyze the reaction. 2 molecules of ATP are generated, thus the net gain of ATP in glycolysis is 2.
• Conversion of phosphoenolpyruvate to pyruvate is an internal oxidation-reduction
• What enzymes catalyzes alcoholic fermentation?

  Pyruvate decarboxylase which removes a CO2 group from pyruvate to make acetaldehyde. Acetaldehyde gets reduced to ethanol by alcohol dehydrogenase which oxidizes NADH to NAD+
• What is the goal of fermentation?

  To oxidize NADH to NAD+ to replenish NAD+ stores so glycolysis may continue. NAD+ is used up when glyceraldehyde 3-phosphate dehydrogenase reduces NAD+ to NADH to create 1,3-BPG from glyceraldehyde 3-phosphate
• In fermentation, organic molecules are the donors and acceptors of elections, not O2 due to anerobic conditions
• What enzyme is present in lactic acid fermentation?

  Lactate dehydrogenase
• In metabolic pathways, enzymes catalyzing irreversible reactions are potential sites of control in the pathway. What enzymes regulate glycolysis?


  hexokinase, phosphofructose kinase, and pyruvate kinase. These enzymes become more or less active in response ti allosteric effectors or covalent modification
• Phosphofructokinase is the most important control site in the mammalian glycolytic pathway. How is it regulated?

  High levels of ATP allosterically inhibit phosphofructokinase by binding to a regulatory site and lowering the enzyme's affinity for fructose 6-phosphate. AMP reverses the inhibitory action of ATP and competes with ATP for the binding site but does not inhibit the enzyme. Therefore, the activity of the phosphofructokinase increases with a lower ATP/AMP ratio.

  A decrease in pH increases the inhibitory effect of ATP (useful in protecting muscles from excess lactic acid)
• Why does AMP but not ADP stimulate the activity of phosphofructose kinase?

  When ATP is being used rapidly, it can also be reformed rapidly from ADP thus ADP cannot be a signal for the low-energy state, while AMP must be.
• Regulation of hexokinase

  Hexokinase is inhibited by its product, glucose 6-phosphate. High concentrations of glucose 6-phosphate signals that the cell no longer requires glucose for energy.
• how does hexokinase and phosphofructokinase communicate with each other?

  When phosphofructokinase is inhibited, the concentration of fructose 6-phosphate rises, and since glucose 6-phosphate is in equilibrium with fructose 6-phosphate, glucose 6-phosphate levels also rise inhibiting hexokinase.