Biochem Lect 26: Glycolysis Part II

2 phases of glycolysis (overall summary)

Phase 1 = glucose → 2 G3P (-2 ATP)

Phase 2 = 2 G3P → 2 pyruvates (+4 ATP, +2 NADH)

Overall Summary for Phase II

everything is doubled!!!

6) G3P → 1,3BPG (“six, redox”)

7) 1,3BPG → 3PG

8) 3PG → 2PG = “3 to 2”

9) 2PG → PEP

10) PEP → Pyruvate

Phase II enzymes

6) G3P dehydrogenase

7) Phosphoglycerate kinase

8) Phosphoglycerate mutase

9) Enolase

10) Pyruvate kinase

Phase II Extra (total)

6) +2 NADH

7) +2 ATP

8)

9)

10) +4 ATP (regulated/-ΔG/irreversible)

Reaction 6

Redox reaction*

Glyceraldehyde 3 Phosphate (G3P) → 1,3 Biphosphoglycerate (1,3 BPG)

Enzyme = G3P dehydrogenase

+1 NADH

Step 6 mechanism (3 steps + notes)

• involves covalent catalysis

• inorganic phosphate provides P

1) SH of Cys attacks G3P → thiohemiacetal

2) thiohemiacetal loses H+ to NAD+ → thioester

3) phosphorolysis (Pi attacks and cleaves SH)

Reaction 7

Small ATP Formation

1,3 Biphosphoglycerate (1,3 BPG) → 3 Phosphoglycerate (3PG)

Enzyme = Phosphoglycerate kinase

+1 ATP

-

spontaneous reaction (-ΔG)!

Step 7 involves _______ phosphorylation

substrate level

-

phosphate is added from substrate to create ATP

phosphoglycerate kinase induced fit

nothing rlly to note tbh

What is 2,3 BPG?

regulator of hemoglobin (found a lot in erythrocytes)

-

made from 1,3 BPG

2,3 BPG is made from _____ and _____

1,3BPG and 3PG

1,3 BPG ←→ 2,3BPG involves what enzyme?

BPG mutase

2,3 BPG ←→ 3PG involves what enzyme?

2,3 BPG phosphatase

summary of equilibrium between 1,3 BPG / 2,3 BPG / 3 PG (view image)

1,3 BPG ←→ 2,3 BPG requires ____

3PG 

-

intermolecular switch

Reaction 8

“3 to 2”

3 Phosphoglycerate (3PG) → 2 Phosphoglycerate (2PG)

Enzyme = Phosphoglycerate mutase

Is step 8 intramolecular or intermolecular?

intramolecular

(Step 7 was intermolecular)

What residue is in the active site of PG mutase?

Histidine

Step 8 mechanism (3 steps) *

1) histidine gets phosphorylated by 2,3 BPG

2) phosphohistidine adds phosphate to C2 

3) phosphohistidine removes phosphate from C3

Phosphohistidine is…

a phosphorylated histidine intermediate

Zelda Rose showed that a little _____ is needed to ________

2,3 BPG; phosphorylate His

Reaction 9

2PG → PEP

2 Phosphoglycerate (2PG) → Phosphoenolpyruvate (PEP)

Enzyme = Enolase

Step 8 purpose

make high energy phosphate (enol) for ATP synthesis (step 10)

enolase structure

assymetric dimer

one subunit holds 2PG, other holds PEP

Reaction 10

ATP Formation

PEP → Pyruvate

Enzyme = Pyruvate Kinase

+2 ATP

very spontaneous reaction (- large ΔG)

Step 10 regulation (4+1)

• ATP ______/AMP ______

• acetyl CoA ______

• F1,6BP ______

• alanine ______ (related to pyruvate)

• liver PK is regulated by _______________

• ATP inhibits/AMP activates

• acetyl CoA inhibits

• F1,6BP activates

• alanine inhibits (related to pyruvate)

• liver PK is regulated by covalent modification (phosphorylation)

pyruvate’s _______ is spontaneous and helps reaction to be thermodynamically favorable

keto-enol tautomerism

in liver, high blood glucose → insulin → pyruvate kinase is ______ → more/less active

dephosphorylated, more

in liver, low blood glucose → glucagon → pyruvate kinase is ______ → more/less active

phosphorylated, less

summary of liver PK regulation (image)

Metabolic fates of NADH (2)

1) aerobic → NADH oxidized in ETC => more ATP

2) anaerobic → NADH oxidized by lactate dehydrogenase (LDH) => more NAD+ (for more glycolysis)

Metabolic fates of Pyruvate (3) - review

1) aerobic = TCA cycle → acetyl-CoA

2) anaerobic = lactic acid fermentation (ex: muscle) → lactate

3) anaerobic in yeast = alcoholic fermentation → ethanol + CO₂

lactic acid bacteria produce…

lactate (yoghurt, sauerkraut…)

Main difference between glucose → pyruvate vs glucose → lactate

no NADH produced when making lactate

Which steps of glycolysis are regulated?

1, 3, 10

Which steps of glycolysis is the “payout” (ATP produced)?

7, 10

Fermentation

ATP produced where organic molecules are electron donor/acceptors

-

(unlike respiration where O2 is electron acceptor)

-

ex: acetone, butanol

The Entner-Doudoroff pathway

tequila fermentation

-

makes more ethanol and at higher concentration

Alternatives to glucose for glycolysis (3 main)

1) Galactose (via Leloir Pathway)

2) Glycerol

3) Lactose

Leloir Pathway

use galactose instead of glucose for glycolysis

-

galactose → gluc 6P

3 main steps of Leloir pathway

1) galactose → galactose 1P

2) galactose 1P → glucose 1P

3) glucose 1P → glucose 6P

How does step 2 of Leloir pathway occur?

UDP glucose and 1P galactose swap phosphates => UDP galactose and 1P glucose 

In step 2 of Leloir pathway, which enzyme swaps UDP?

galactose 1P uridyl transferase

In step 2 of Leloir pathway, which enzyme switches UDP galactose with UDP glucose?

UDP galactose 4 epimerase

Glycerol can also enter glycolysis as ____ in 2 steps:

DHAP

1) phosphorylate glycerol

2) redox → DHAP + NADH

Glycerol → DHAP requires 2 enzymes:

1) glycerol kinase

2) glycerol phosphate dehydrogenase

Lactose can be converted to glucose via…

cleavage

-

lactose = disaccharide of galactose + glucose

____ synthesizes lactose and _____ breaks down lactose

lactase synthase, lactase