boichem: module 4

what are the 2 phases of glycolysis

1. energy investment

   1. glucose → G6P

   2. G6P → F6P

   3. FBP →

   4. GAP + DHAP

2. energy payoff

   1. GAP → 1,3-BPG

   2. 1,3-BPG → 3PG

   3. 3PG → 2PG

   4. 2PG → PEP

   5. PEP → pyruvate

glycolysis: phase 1

• energy investment

• goal: prepare glucose for cleavage into two 3-carbon units

• what happens:

  • glucose is phosphorylated (trapped in cell)

  • isomerized to fructose

  • phosphorylated again (committed step)

  • cleaved into two triose phosphates

  • interconverted to same form (G-3-P)

• energy cost: 2 ATP invested

glycolysis: phase 2

• energy payoff (steps 6-10)

• goal: produce ATP and NADH while converting G-3-P to pyruvate

• what happens:

  • G3P is oxidized → NADH formed

  • ATP generated (substrate-level phosphorylation)

  • dehydration forms high energy PEP

  • ATP generated from PEP → pyruvate formed

• energy gain: 2 net ATP produced, 2 NADH

• net yield: 2 pyruvate (3 carbons each) + 2 ATP (net) + 2 NADH

step 1: hexokinase reaction

• reaction: glucose + ATP → glucose-6-phosphate + ADP +Pi

• first ATP investment

• phosphorylation traps glucose in cell

• this reaction is irreversible under intracellular conditions

• metabolic activation or priming of glucose

• broad substrate specificity (can phosphorylate other hexoses)

why does phosphorylation of glucose by hexokinase matter

• Glucose:

  • uncharged can cross membrane via GLUT transporters

• glucose-6-p:

  • negatively charged

  • CANNOT cross plasma membrane

• kinetic properties:

  • Km for glucose ~ 0.1mM

  • [glucose] in the cell: ~4mM

  • this means the enzyme operates near Vmax (always active when glucose is present)

  • regulation: allosteric inhibition by the product (G-6-P)

what are the tissue specific isoforms of hexokinase

• isoform I:

  • tissue expression: brain, RBCs

  • regulation: strong G6P inhibiton; high glucose affinity

• isoform II:

  • tissue expression: muscle, adipose tissue

  • regulation: G6P inhibition; insulin-responsive

• isoform III:

  • tissue expression: various tissue

  • regulation: similar to I and II; less well characterized

• isoform IV (glucokinase):

  • tissue expression: liver, pancreatic beta-cells

  • regulation: not inhibited by G6P

  • regulated by glucokinase regulatory protein (GKRP) and insulin

where do hexokinase and glucokinase target

• hexokinase:

  • muscles

  • high affinity (Km 0.1mM)

  • inhibited by G6P

• glucokinase:

  • liver/pancreas

  • low affinity (Km 10mM)

  • acts as a “glucose sensor”

how does glucokinase act as the glucose sensor

• high Km (~10mM) - only active at HIGH [glucose]

• NOT inhibited by G6P

• regulated by glucokinase regulatory protein (GKRP)

• role in pancreatic B-cells

  • glucose enters via GLUT2 (high Km transporter)

  • GK phosphorylates glucose → ↑ Glycolysis/TCA/ETC

  • ↑ ATP/ADP ratio

  • ATP sensitive K+ channels CLOSE

  • membrane depolarizes

  • voltage-gated Ca2+ channels OPEN

  • Ca2+ influx triggers insulin vesicle EXOCYTOSIS

how is glucagon secreted by alpha cells

1. during hypoglycemia [glucose] falls

2. lower ATP/ADP ratio

3. ATP-sensitive K+ channels CLOSE

4. intracellular K+ rises

5. membrane depolarizes

6. voltage-gated Ca2+ channels OPEN

7. Ca2+ influx triggers glucagon vesicle EXOCYTOSIS

how is G6P a key branch point

• its fate depends on a cellular needs

step 2: phosphoglucose isomerase reaction

• reaction:

  • G6P → F6P (reversible)

    • ΔG°' = +1.67 kJ/mol

  • Why this step?

    • converts aldose to ketose

    • positions C1 for phosphorylation in step 3

    • prepares molecule for symmetric cleavage in step 4

phosphoglucose isomerase mechanism

• opening of the pyranose ring

• proton abstraction leading to enediol formation

• proton addition to the double bond followed by ring closure