7 Glycolysis 20140920
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Glucose Metabolism
C6H12O6
Complete Oxidation of Glucose
+ 6O2 6CO2 + 6H2O
Glucose Metabolism
• Central position in metabolism • A good cellular fuel (- 2,840 kJ/mol for complete oxidation) • Released from high molecular weight polymers • Production of ATP • Supplies metabolic intermediates for biosynthesis
Glucose
Fates of glucose in animals and plants
• Storage
• Glycolysis: oxidation to 3-carbon compounds to provide ATP and metabolic intermediates
• Pentose phosphate pathway - oxidation to yield ribose-5-phosphate and NADPH
Synthesis of glucose
• Gluconeogenesis
• Photosynthesis
Glycolysis - A series of enzyme-catalyzed reactions in a pathway
- Overall conversion:
- Generation of 2 ATPs (net gain)
- These reactions are the way in which carbohydrate is broken down into smaller units. - Anaerobic metabolism: pyruvate is converted to lactate or ethanol (fermentation) - Aerobic metabolism: pyruvate is fed into the TCA cycle where it is fully metabolized to carbon dioxide releasing more ATPs
Hexose stage: (Preparatory phase)
Triose stage: (Payoff phase)
The ten steps of glycolysis
Hexose stage: 1 to 4 Triose stage: 5 to 10
(1) Hexokinase: phosphorylation of glucose
Hexokinases - Hexose kinases - Four isoenzymes isolated from mammalian liver - Hexokinase I, II, III: Km for glucose = 10-6 – 10-4 M - Hexokinase IV = Glucokinase: Km for glucose = 10-2 M (more active at high blood glucose conc.)
(2) Conversion of G-6-P to F-6-P:
(aldose-ketose isomerization)
(3) Transfer of a second phosphoryl group from ATP to F-6-P
- The first committed step in glycolysis
(β-D-fructofuranose form)
PFK-1
Glucose Glucose 6-P Fructose 6-P
Fructose 1,6-bisP
PFK-1
Glycolysis only
other pathways
other pathways other pathways
(4) Cleavage of C3-C4 bond - Production of two triose phosphates
- Unfavorable under standard condition (G > 0) - Actual free energy change ~ 0, thus a near-equilibrium reaction - Rapid consumption of DHAP and G3P “pulls” the reaction forward
Aldolases - Class I: plants and animals - Class II: microorganisms - Classes I and II are not structurally related - An example of “convergent evolution”
Mechanism of F1,6-bisP cleavage by aldolases:
X – an electron withdrawing group - Lysine amino group in Class I - Zn2+ cofactor in class II
(5) Conversion of DHAP to G3P
- Enzyme mechanism: general acid-base catalysis (see lecture notes on enzyme mechanism)
- Only G3P continues in the glycolytic pathway - Which direction of the reaction is favored?
Fate of carbon atoms from the hexose stage to the triose stage:
(6) Formation of a “high-energy” compound: 1,3-Bisphosphoglycerate
Acid anhydride linkage
- Beginning of the recovery of energy from triose phosphates - Oxidation-reduction reaction catalyzed by a dehydrogenase (why is it called a
dehydrogenase?) - Phosphorylation reaction (how is it different from the earlier kinase reactions?)
(7) Generation of ATP - 1,3-BPG hydrolysis releases large amount of energy - Substrate level phosphorylation
- Stabilization of product by resonance hybrid formation:
- G3P dehydrogenase and PG kinase associate to form a complex: efficient channeling of 1,3-bisphosphoglycerate
Arsenate poisoning - AsO4
3- replaces inorganic phosphate (PO43-) in phosphoryl transfer
reactions
Glyceraldehyde 3-P +
AsO43-
+ NAD+
Glyceraldehyde 3-P dehydrogenase
(No 1,3-pisphosphoglycerate production)
(8) Intramolecular phosphoryl group transfer
Phosphoglycerate mutase
Mutase: an isomerase that catalyzes the intramolecular shifting of a chemical group
2
3
2
3
Phosphoglycerate kinase: mechanism
(9) Dehydration to an energy-rich compound, PEP
(PEP)
(10) Generation of ATP
- PEP: high-energy phosphate compound
- Tautomerization stabilizes products of hydrolysis
- Substrate level phosphorylation
- Glycolysis finally turns a profit!
tautomerization
(keto form) (enol form)
Overall process of glycolysis
Glucose + 2ATP + 2NAD+ + 4ADP + 2Pi → 2 pyruvate + 2ADP + 2NADH + 2H+ + 4ATP + 2H2O Simplifying the equation will give you: Glucose + 2NAD+ + 2ADP + 2Pi → 2 pyruvate + 2NADH + 2H+ + 2ATP + 2H2O
The fate of pyruvate
Metabolism of pyruvate to ethanol
- Regeneration of NAD+ for glycolysis
Summary of glycolysis and ethanol formation
Reduction of pyruvate to lactate
- Regeneration of NAD+ for glycolysis - Anaerobic bacteria - Mammals (skeletal muscles during vigorous exercise)
Summary of glycolysis and lactate formation
Other sugars can enter glycolysis
Entry of fructose into glycolysis through fructokinase
Conversion of mannose to fructose 6-phosphate
Conversion of galactose to glucose 6-phosphate
Regulation of glycolysis
1. Regulation of hexose transporters
- Membrane-embedded transporters (e.g. GLUT family transporters) - Stimulation of cellular glucose uptake by insulin
(Skeletal and heart muscle cells, Adipocytes)
2. Regulation at 3 irreversible steps
(a) Hexokinase
Glucose + ATP Glucose 6-phosphate (G-6-P) + ADP G-6-P - Allosteric inhibitor of hexokinases I, II, III (found in muscles)
Glucokinase - Not inhibited by G-6-P - the major hexokinase in liver - Converts glucose to G-6-P in liver after a meal - G-6-P is used for glycogen synthesis when glucose is sufficient in other tissues - High Km value for glucose - Never saturated with glucose - Activity increases with increasing concentration of available glucose
(b) Phosphofructokinase-1 (PFK-1)
Fructose 6-phosphate + ATP Fructose 1,6-bisphosphate + ADP
ATP - a substrate and an allosteric inhibitor AMP and ADP – allosteric activators
ATP reduces the affinity of PFK-1 for F 6-P
AMP relieves the inhibition by ATP
Citrate – feedback inhibitor; a TCA cycle intermediate - high conc indicates that TCA cycle is blocked
Fructose 2, 6-bisphosphate – an allosteric activator of PFK-1
Glucagon
Low blood glucose
In liver cells:
+
PFK-1 less active
(Phosphofructokinase-2)
(c) Pyruvate kinase
Phosphoenolpyruvate (PEP) + ADP Pyruvate + ATP
Fructose 1,6-bisphosphate (F1,6BP) – an allosteric activator
ATP – a strong allosteric inhibitor
PFK-1 , F1,6BP , Pyruvate kinase Thus, PFK-1 activation leads to subsequent pyruvate kinase activation - Feed-forward activation
The Entner-Doudoroff Pathway in Bacteria
- No PFK-1 enzyme in bacteria - No fructose 1,6-bisphosphate
formation from glucose - Generates fewer ATP than glycolysis,
why? - Earliest pathway for glucose
degradation
PFK-1 (not in some bacteria)
fructose 1,6-bisphosphate
fructose 6-phosphate
Two products in this pathway: Pyruvate (end product of glycolysis) Glyceraldehyde 3-phosphate
Glycolysis (triose phase)
Pyruvate
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