Transcript U4L21 fuel oxidation - The University of Sydney

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Fatty acid oxidation
• Also called beta-oxidation
– Because the action occurs on the beta-carbon
atom
Fatty acid oxidation
• Requires tissues to have mitochondria
• Reciprocally regulated with glucose
oxidation
– Fatty acid oxidation inhibits glucose oxidation
• Consumes a lot of FAD, NAD, CoA
– Availability of cofactors is important
Transport of FA
Fatty acid binding protein
Albumin
Transport of FA
• FA needs to be transported from blood into tissues
• FA is carried in blood on albumin
– Several binding sites for FA
• There are specific transporters for FA
– CD36 moves to the cell surface whenever there is a
need to take up FA at a rapid rate
• FA is carried on FABP (fatty acid binding protein) in
cytoplasm
Trapping of FA
• FA is trapped by attaching it to CoA
• This also ‘activates’ the fatty acid (‘tags’ the FA)
• Requires quite a lot of energy,
– ATP is not converted into ADP, but AMP
Transport of FA: Mitochondria
FA-CoA is oxidized
Example: 16C FA-CoA
• 7 NADH & 7 FADH2 are produced…. NAD & FAD needed
• 8 acetyl CoA produced…. CoA needed
Cofactor Availability
• NAD, FAD and CoA
– All needed to keep FA oxidation going
• How are these carriers regenerated?
– CoA
• By entry of acetyl CoA into Krebs Cycle
– NAD/FAD
• By giving cargo to electron transport chain
Rate Limiting Enzymes
• The slowest enzyme in the metabolic
pathway determines the overall speed
– Rate-limiting step (RLS)
– Flux generating step
• Key points of regulation
Enzyme kinetics 
•
Vmax
•
Rate
½ Vmax
Km
S1
[substrate]
S2
At high [substrate],
minor changes in
[substrate] will not
affect the rate of
reaction
Doubling or
halving the [S] isn’t
even going to
affect the rate
Redfern Station Analogy
•
Imagine the station at peak hour with just
one barrier operating
– This gate will soon become ‘saturated’ with
people
– Increasing the number of people doesn’t
increase the rate
– It is the ‘rate limiting’ step
– The point which determines the overall rate at
which people get to Uni
Changing the Flux
•
There are 3 major ways to regulate this
(and metabolic!) pathways
– Change the intrinsic activity of the step
•
Make ticket-reading & gate-opening happen faster
– Make more gates open
•
•
•
Switch them from being ‘off’ to ‘on’
Or change the direction from ‘in’ to out
Or bring in a set of gates when you need them
– Make and destroy gates according to need
•
Seems crazy!
Properties of RLS
• Irreversible
– Need alternative enzymes to ‘go back’
– Not ‘equilibrium’ under physiological
conditions
– “Committed steps”
• Saturated with substrate
– Low Km or [S] >> Km
– Working at Vmax
RLS in FA ox?
•
•
•
•
Availability of fatty acids?
Cell membrane transport & Trapping?
Mitochondrial transport? Carnitine
Oxidation?
– Activity of enzymes
• Co-factor availability?
• Does it depend on the circumstances?
Glycolysis
•
•
•
•
•
•
Uses carbohydrate (glucose)
Wholly cytosolic
All cells of the body
No requirement for oxygen
Very, very fast
Very inefficient
Glucose
Glucose Uptake
P
hexokinase
glucose
blood
glucose
Using ATP
glucose
6-phosphate
cytoplasm
Uptake facilitated by Glucose transporters (GLUTs)
•GLUT-1 present in all cells all the time
•GLUT-4 muscle and adipose tissue (the insulin sensitive tissues)
•GLUT-2 liver and pancreas (blood glucose regulating tissues)
Early Glycolysis
Investment of energy giving a biphosphorylated symmetrical sugar
P
P
Phosphofructokinase
glucose
6-phosphate
PFK
fructose
6-phosphate
P
P
Using ATP
fructose
1,6-bisphosphate
Splitting to give two 3-carbon molecules
P
P
Two molecules of 3-carbon sugar phosphates
Return Phase
Remember two 3-carbon
molecules go down the pathway
P
Bring in phosphate
Oxidize with NAD
P
P
Super energy molecule!
Recoup some ATP
P
Recoup some ATP
pyruvate
Overview
• Total yield is 2 ATP per glucose
– And two pyruvate
– And two NADH
• Need to regenerate NAD
• Fate of the pyruvate
– Aerobic
– Anaerobic
Completing Glycolysis
• More ATP from oxidation of pyruvate
– Need to transport into mitochondria
– Oxidize with pyruvate dehydrogenase
• Need to reoxidise NADH
– To maintain the supply of NAD
– Shuttle systems available
• To send NADH electrons/hydrogens into matrix
– Lactate production
– In yeast, alcohol production
– Latter two keep everything cytosolic
Regulation
• Most points reversible
• Focus on three steps
– Hexokinase (G  G6P)
• Mainly feedback inhibition from G6P
– Phosphofructokinase (F6P  F6BP)
• Strongly affected by ATP/ADP levels
• But mainly via AMP levels
– Pyruvate kinase (last step)
• ATP/ADP important
Energy Charge
• Large changes in ATP not desirable
– Keep ATP at 5 mM
• Adenylate kinase
– Translates small change in ATP to large
relative change in AMP
– 2ADP  ATP + AMP
• Ratio of adenine nucleotide concentrations
often called ‘energy charge’
• Strong stimulation of PFK
Energy Charge
Integration of Catabolism
FA
BETA-OXIDATION
“CARNITINE”
FA-CoA
CD36
ac-CoA
ac-CoA
ac-CoA
FA-CoA
ac-CoA
ac-CoA
PDH
pyruvate
GLUT-4
glucose
citrate
GLYCOLYSIS
G6P
OAA
pyruvate
KREBS CYCLE
PHOSPHORYLASE
CO2
glycogen
CO2
IC
ICDH
2OG
OGDH
Krebs Cycle
• TCA cycle, Citric Acid cycle
• Substrate is acetyl CoA
– Fatty acid oxidation and/or glucose oxidation
• Overall strategy
–
–
–
–
Completely oxidize acetate carbons to CO2
Produce lots of NADH, FADH2, even an ATP
Perform the reaction on a carrier molecule
Regenerate the carrier
Regulation
• Krebs cycle activity is controlled early on
– At isocitrate dehydrogenase (ICDH)
– alpha-ketoglutarate dehydrogenase (OGDH)
• ICDH and OGDH are stimulated by rise in Ca2+
– Such as is found during exercise
• ICDH & OGDH are also sensitive to NAD levels
– Activity is dependent on availability of NAD
Important Features
• During the cycle
– 2 carbon atoms come in, 2 carbon atoms released
• Generates
– 3 NADH, 1 reduced FAD plus a GTP
– Each NADH gives 2.5 ATP in oxidative
phosphorylation
– Each FADH2 gives 1.5…
– So with the GTP, that’s about 10 ATP per acetate
• Oxaloacetate is not ‘net’ consumed in the cycle
– acts as carrier