Week 1B: Metabolism Basics, Organs and metabolic pathways, glycolysis and pentose phosphate pathway, TCA cycle Flashcards
HC03-06
HC03: Energy methods out of glycolysis
-NADH
-ATP
Requirement of free energy for:
-Mechanical work: cellular transport, muscle contraction
-Active transport of molecules and ions
-Synthesis of macromolecules from building blocks
-Thermogenesis
Energy is obtained from the … of food
oxidation
Metabolism consists of … pathways of chemical reactions
Interconnected
Anabolism
Synthesis of complex molecules
- useful energy + simple precursors > complex molecules
- Gluconeogenesis of glucose from pyruvate or lactate for example
Catabolism
Breakdown
> Carbohydrates and fats to CO2, H2O and useful energy
> Glycolysis of glucose
Metabolism is regulated, why?
You don’t want to make something and break it down simultaneously
-Glycolysis yields 2 ATP
-Gluconeogenesis costs 6 ATP
> organize the fluxes
The currency of free energy
ATP: adenosine triphosphate
ATP structure
Adenine, ribose and 3 phosphates
How many energy rich bonds does ATP have and how are they called?
Two phosphoanhydride bonds
> Gamma and beta phosphates are energy rich bond
> the outer bond (gamma) is between two phosphates (both negatively charged)
> alpha bond not between two phosphates, lower energy
ATP can be converted to … to release energy
ADP, or AMP
dG and entropy formula
dG = dH - T*dS
dG: delta Gibbs free energy
dH: delta enthalpy (heat content)
dS: entropy: degree of disorder
ATP>ADP+Pi is an … reaction (hydrolysis)
Exergonic (energy released for Gibbs free energy)
What is the dG0’ of ATP > ADP
-30.5 kJ/mole
What if dG0’ is negative?
Spontaneous reaction
> can be coupled to an energetically unfavorable reaction
Why are phosphoanhydride bonds energy rich
-A lot of resonance structures available where the energy can be located
- Force of the negative charges who lay in close proximity
> ATP 4- > outer phosphate 2- and middle and inner 1-
dG0’ of ATP > AMP + PPi. When is this used?
dG0’=-45.6 kJ/mole
> used if the energetically unfavorable reaction has a dG0’ > 30.5 kJ/mole
In which organisms is the ATP-ADP cycle the fundamental mode of energy exchange?
All
Why are phosphoenolpyruvate, 1,3-BPG and creatine phosphate, with a more negative dG0’ for conversion less good as energy currency
These compounds can be used to phosphorylate ATP fro ADP
> you can synthesize ATP without oxidative phosphorylation in mitochondria if higher energy molecules are oxidized. If the currency is a maximal energy carrier, this is not easily possible.
How is it called if high energy compounds are used/oxidized to make ATP? How much of the ATP is made through this mechanism?
Substrate-level phosphorylation (10%)
Creatine usage before exercise
In muscle cells, you make creatine phosphate using ATP before the sport in rest
> During exercise, creatine phosphate can be used to synthesize ATP as early reserve for substrate-level phosphorylation
> for sudden exercise
> oxygen cannot hold up with muscle activity, the oxidative phosphorylation cannot keep up with the use of ATP, creatine supplementation to make ATP with substrate level phosphorylation on short term
Creatine is synthesized in our …
Liver and kidneys
Creatine phosphate + ADP <=> ATP + creatine: enzyme?
Creatine kinase
ADP+ADP <=> ATP + AMP. Explain and enzyme
It costs two ATP to regenerate ATP from AMP. Enzyme is adenylate kinase
> this reaction is used when ATP levels run low in human cells including muscle. Produce ATP from ADP for extra energy
> AMP is useless until regeneration by AMP kinase if there is much AMP
Sources of ATP during exercise
Seconds: ATP reserve and creatine phosphate
Minutes and hours: Anaerobic and aerobic metabolism (ATP regeneration)
In initial seconds after cold start, ATP is regenerated by …. from ADP and creatine phosphate, followed by metabolic pathways
High-phosphoryl transfer
Oxygen uptake during exercise
At start, oxygen uptake increases exponentially, until a steady state is reached
> first minutes exercise: shortage of oxygen
> substrate level phosphorylation is then essential for energy supply
Human muscle fibers
-Type I: slow twitch: long time (hours), aerobically, low power, high density mitochondria, 80msec till peak contraction
-Type IIa: fast twitch a/ intermediate: <30min, middle power, middle density mitochondria, 30 sec till peak
-Type IIb: fast twitch b: High intensity, <5 minutes exercise, low density mitochondria, anaerobically.
> most fibers I, than IIa, than IIb
Human body contains approx … g of ATP, and at rest we need …
100
at rest: 40 ATP a day
> high turnover is required for the relatively small amounts of ATP
ATP used for..
-Motion, without it muscles freeze. To return actin-myosin to relaxed state (crossbridge cycle)
-Active transport: in muscle, Ca2+ ATPase uses ATP for transport Ca2+, needed in contraction
-Biosynthesis
-Signal amplification
Most efficient fuel
Fats
Redox reaction principles
Reduction: gain of electrons
Oxidation: loss of electrons
Electron transfer from reduced compound (oxidation) to the oxidized compound (reduction)
Oxidation states of C1’s go in …
two electron reactions
Most energy (reduced)
Methane CH4 (-4)
- 2 e-
Methanol (hydroxyl) CH3OH (-2)
Formaldehyde (aldehyde, ketone) HC(=O)H (0)
Formic acid (carboxyl) HC(=O)OH (+2)
Carbon dioxide (O=)C(=O) (+4)
Most important fuels
Glucose and fats
> fats more efficient, because the carbon atoms are in a more reduced state
Direct burning of sugars
All Gibbs free energy released as heat, waste
Glucose oxidation in the cell
In small steps by enzymes to tranfer Gibbs free energy to carrier molecules
How many steps for glucose to pyruvate
10
Why does something change upon phosphorylation of a protein
Different charges negative on phosphorylated site
Conformational change after phosphorylation Ca2+ ATPase
Calcium ion flips out (transport over membrane) on the other site because of conformational change of teh ATPase which spans over the membrane > into lumen
> dephosphorylation and reset for next cycle
Oxygen wants to …
take up electrons (acceptor)
> carbon and oxygen next to it: unfortunate, electron gets pulled awat
> carbons have more electrons with just hydrogens next to it
Energy in glycolysis is released in oxidation of an aldehyde (GAP, glyceraldehyde 3-phosphate) to a carboxylic acid (3-phosphoglyceric acid). How?
Through intermediate step, energy of oxidation is first trapped as high potential phosphate group
> two electrons plus H+ ion (hydride ion H-) released and captures by electron carrier NAD+
> GAP + NAD+ + HPO4- > 1,3-BPG (intermediate) + NADH + H+
> 1,3-BPG (1,3-bisphosphoglycerate) + ADP > 3-phosphoglyceric acid (with extra OH at place of phosphorylation for intermediate, carboxyl end) + ATP
Oxidation GAP to yield ATP is an example of
Substrate-level oxidation
When is Gibbs free energy stored in ion gradients over membranes?
Proton gradient, fueled by oxidation of fuels by special proton pumps
> oxidative phosphorylation in mitochondria
> yields 90% of the ATP
Energy extraction from food
1: macromolecules are broken down into small units (stage 1)
2: breakdown to acetyl group of acetyl-CoA, a central metabolite
3: ATP production through the oxidation of the acetyl group of acetyl-CoA
Activated carriers in metabolism
-ATP: phosphate groups high potential
-NADH and FADH2: activated carriers of electrons during oxidation of fuels
-NADPH: activated carrier of electrons for reductive biosynthesis
-Coenzyme A: activated carrier of carbons
Characteristics NADH and NADPH
-Water soluble co-enzymes that carry electrons
-Contain nicotinamide ringwith reactive site at the carbon opposite to the N.
> different R groups in standard structure, in NAD+: H, in NADP+: PO3(2-)
-NADH for oxidative phosphorylation and NADPH for reductive biosynthesis
What do NAD+ and NADP+ accept to be reduced?
A hydride ion: H+ with two e-.
> in addition, a second H+ is split of the molecule which is oxidized by NAD+ and appears in the solvent
Reactive sites FAD+
Two nigle N’s on the FMN (flavine mononucleotide)
Characteristics FAD
In flavoproteins, a FMN or FAD (flavine adenine dinucleotide) are tightly bound co-enzymes: prosthetic group.
> FAD is more flexible than NAD+ and can participate in transfer of single hydrogen atom (electron and proton) or in the transfer of two H atoms.
Reducing FAD
The two hydrogen atoms are transferred to FAD to form FADH2. The two H are added to the free N in the three ring structure
Which electron carrrier can participate in the most diverse set of reactions?
Flavoproteins instead of NAD(P)-dependent enzymes > can catalyze transfer of one or two electrons
Redox enzymes
Oxido-reductases
Which enzymes remove hydrogen atoms?
Dehydrogenases
Function oxidase?
Transfer of only electrons. Uses molecular oxygen O2 as the acceptor of the electrons from the oxidized compound.
Function oxigenase
Oxidizes a compound by adding O2 from molecular oxygen to it
Complex IV function (cytochrome c oxidase)
Transfers electrons to molecular oxygen which is reduced to water
4 Cytochrome c red + 4H+ + O2 > 4 Cytochrome c ox + 2 H2O
Number of valency electrons in O2 an H2O
Octa valency rule but for hydrogen just two (2 stripes, 4 electrons)
O2: 2 * 6 = 12
H2O: 8 (2 stripes = 4 from O, 1 stripe = 2 per H)
Successive one-electron reductions of molecular oxygens
Oxygen (O2)
> Superoxide anion (O2-)
> Hydrogen peroxide (H2O2) (addition 2 H+)
> Hydroxyl radical (OH) + hydroxide (OH-)
> 2 H2O water (addition 2 H+)
The three reactive oxygen species (ROS)
Three intermediates in oxygen reduction to water
> Superoxide (O2-)
> Hydrogen peroxide (H2O2)
> Hydroxyl radical (OH)
Most dangerous ROS
Hydroxyl radical (*OH), the most reactive free radical, initiates oxidative destruction of biomolecules
What group is transferred by a dehydrogenase?
A hydride ion (two electrons and H+) > oxidation.
Lactate dehydrogenase reaction
Lactate > pyruvate
Using NAD+ to accepts the hydride ion
Effect of high levels of NADH on gluconeogenesis
High levels of NADH inhibit the oxidation of lactate to pyruvate in the liver
> reduction liver pyruvate to lactate
> Hypoglycemia and lactate acidosis
Cori cycle
In muscle:
Glucose > pyruvate > lactate (yields 2 ATP)
to blood to liver
Lactate > pyruvate > glucose (costs 6 ATP)
glucose to blood to muscle
The TCA ccle has two simple oxidation reaction catalyzed by
-Succinate dehydroenase (succinate > fumarate)
-Malate dehydrogenase (malate > oxaloacetate)
> yield NADH
Oxidative decarboxylation reactions in TCA cycle with oxidation which also yield NADH (also oxidation)
-Isocitrate dehydrogenase (isocitrate + NAD+ > a-ketoglutarate + CO2 + NADH + H+)
-a-ketoglutarate dehydrogenase (a-ketoglutarate + NAD+ + CoA > succinyl-CoA + CO2 + NADH + H+)
How many electrons for reduction oxygen to water?
4 electrons
Difference oxygen and carbon reaction
Oxygen deals with one electron and carbon only with two (often hydride ion)
Which compounds are needed to bind ROS in oxidation O2?
Transition metals
CoA reactive group and carry of acetyl-CoA
The HS- which can form a high energy thioester bond with an carboxyl group.
> acetyl-CoA carries an activated acetyl group.
dG0’ of acetyl-CoA + H2O > acetate + CoA
-31.4 kJ/mole
> high potential thioester bond: -C(=O)-S-CoA
> costs ATP > AMP + PPi (so 2 ATP) to form because 45.6>31.4>30.5 kJ/mole
When is the released energy of thioester bond in CoA carrier used
For example in reductive biosynthesis of cholesterol
> HMG-CoA + 2 NADPH + 2 H+ > mevaonate + 2 NADP+ + CoA (HMG-CoA reductase, committed step)
H05: Carbohydrate metabolism catabolism
Glucose > energy and CO2 and H2O
-Glycolysis
-Pyruvate oxidation (or reduction to lactate, anearobic glycolysis)
-Citric acid cycle
-Electric transport / respiratory chain
-ATP synthesis
Respiratory chain + ATP synthesis =
oxidative phosphorylation
HC04: Metabolism is regulated through control of (3):
-Amount of enzymes
-Catalytic activities of enzymes (hormonal control, allosteric control)
-Accessibility of substrates (compartmentilization)
Energy intake vs expenditure
-Intake through diet: fat, protein, carbohydrate, alcohol
-Expend through thermogenesis, physical activity and basal metabolic rate
Food is broken down to acetyl-CoA. Fates acetyl-CoA and pyruvate
Glucose <=> pyruvate
Pyruvate > (irreversible reaction) acetyl-CoA
Acetyl-CoA
> CO2 (irreversible, oxidation to CO2, TCA cycle for energy)
<=> lipids (storage as fats)
Reaction to insulin in glucose uptake
Increased uptake glucose by muscle cells and adipocytes
Which organ is primarily responsible for maintaining blood glucose levels?
The liver
States of energy
Fed state: insulin rules, anabolic
Fasted state: glucagon rules, catabolic
> pathways directed in liver by hormones
Endocrine pancreas cells
-Alpha cells: synthesize insulin
-Beta cells: synthesize glucagon
-Insulin inhibits the alpha cells from making glucagon
> in islets of Langerhans
Exocrine pancreas makes
Digestive enzymes
Diabetes mellitus type 1 cause
Autoimmune response against own pancreatic beta cell > low insulin, high glucagon (no inhibition, even when hyperglycemia)
If the blood glucose is high than
The beta cells secrete insulin
Insulin effects
It stimulates anabolic anabolism of glycogen and fatty acids, but inhibits gluconeogenesis (because gluconeogenesis in fasted state)
When fuel for energy
Always but not in the fed state when there is enough ATP, storage to fatty acids or glycogen
Main consumers glucose
Brain and erythrocytes
When hypoglycemia
Blood glucose «4.5 mM
Glucose metabolism in fed state
Glucose from gut to portal vein and insulin secretion by pancreas to liver, adipocytes, muscle and all tissues as well.
> Liver converts a part of the glucose to glycogen
> Tissues take up glucose for fuel or for biosynthesis: brain, adipocytes, erythrocytes, muscle cells
> Muscle cells make glycogen
Glucose can exit the liver after activation to glucose-6-phosphate because it expresses the enzyme
G6Pase: Glucose-6-phosphatase
Glucose metabolism in fasted state
Alpha cells in pancreas secrete glucagon
> Glycogen breakdown in liver and transport to the brain and erythrocytes via blood.
> Blood has no mitochondria: anaerobic glycolysis and transfer lactate to liver for conversion to glucose
Fat metabolism in fed state
Glucose and amino acids from gut to portal vein to liver
> Fat from gut through enterocytes and as chylomicrons through lymphatics to blood and TAGs are taken up by muscle cells and adipocytes as FAs (LPL activity)
> secretion insulin
> Liver: glucose to glycogen, and glucose and aminoacids acetyl-CoA to use acetyl-CoA to make fatty acids
> Fatty acids in liver used to make TAGs: transport through VLDL to blood for periphery.
>Muscle uses fatty acids for energy or storage as TAG
> adipocytes store FAs in TAGs
Fat metabolism in fasted state
Glucagon rules
> Liver: glucose to blood to brain and erythrocytes
> breakdown TAGs in adipocytes to FAs, and transport to liver via blood. Liver converts it to acetyl-CoA and uses it for energy
> Muscles take up FAs from blood and convert it to acetyl-CoA for oxidation for energy
Amino acid metabolism in fasted state
glucagon rules
> breakdown proteins to amino acids in gut > to liver via portal vein
> conversion amino acids to glucose in liver for brain and erythrocytes
Precursors gluconeogenesis (in liver) in fasted state to provide glucose to brain and erythrocytes mainly
Amino acids from dietary proteins, lactate from erythrocytes, glycerol from lipolysis in adipocytes.
Central metabolite which interconnects the glucose, fat and amino acid metabolism
Acetyl-CoA
Metabolism in untreated T1DM
No insulin, glucagon rules
- GLUT4 not upregulated: no uptake glucose by adipocytes and muscle cells, just erythrocytes and brain. And glycogen breakdown even when uptake glucose and amino acid conversion to glucose > glucose accumulation
- Protein breakdown in muscle and alanine transport to liver and making glucose there
-Glucagon promotes lipolysis in adipocytes: FAs accumulate in blood (FFAs to serum albumin)
-FAs in liver converted to VLDL with TAGs (accumulate in blood) and ketone bodies (accumulate in blood)
Increasing activation of muscle fibers with increasing force
Type I > Type IIa > Type IIb
- Type I fiber takes the largest burden during light exercise, this decreases during intenser exercise (also more used, but relatively less because also other fibers used now)
Energy sources during exercise
-Increasing efforts: use of blood glucose and muscle glycogen increases while use of muscle fat and blood free fatty acids decrease.
> maximum effor: glycogen is the preferred energy source
HC05: Glycolysis entails the oxidation of .. to
glucose to 2 pyruvate which yields NADH and ATP
There is …. ATP transport across mitochondrial membrane
Coupled antiparallel transport
Expression different GLUT transporters and glucokinase/hexokinase in tissues
Gut: GLUT2
Pancreas beta cells: GLUT2 and GK
Liver: GLUT2 and GK
Brain: GLUT3 and HK
Erythrocytes: GLUT1 and HK
Muscle and adipocytes: GLUT4 and HK
Km and character of GLUT2
GLUT2 for high concentrations glucose in lumen gut import for transport to liver
> expressed by gut, pancreas and liver
> high Km, low affinity (20 mM)
> a lot of the glucose will pass the liver via GLUT2 to the blood stream
Where GLUT1/3 expression, and characteristics
In all mammalian tissues
> For basal glucose uprake
> low Km (1 mM), high affinity
> in brain much expression GLUT3: high affinity, much glucose needed, just like GLUT1 for erythrocytes (much glucose needed, only glycolysis)
GLUT4
In muscle cells and adipose cells: high Km, low affinity (5 mM) and insulin-dependent
> glucose uptake in diabetics patients after insulin injection
> only when insulin rules, in fed state
> the Km is 5 mM, equal to the blood glucose 5 mM. For GLUT1/3 Km 1 mM < 5 mM
Why glucose transporters?
Glucose is hydrophilic and cannot pass the membrane
Sodium glucose linked transporter 1 (SGLT1)
Facilitates active transport of glucose over PM in the enterocytes of the gut > low sodium concentration in cytosol and high in lumen small intestine > cotransport glucose and sodium
- costs ATP hydrolysis to restore sodium balance (low inside cell) with a Na-K ATPase on the basal membrane
-Glucose to blood from intestinal cell with concentration gradient through glucose permease
- active transport makes sure that all glucose is taken up, so the uptake is not completely dependent on the high affinity GLUT2.
Pyruvate fates for energy in exercizing muscle cells
-Last seconds of sprint: low O2, pyruvate reduction to lactate
-Normal: oxidation pyruvate in mitochondria after decarboxylation to acetyl-CoA
First stage glycolysis
Investment: 2 ATP used
> activation glucose to keep it inside the cell
> Hexokinase enzyme phosphorylates glucose to glucose-6-phosphate (cannot pass PM)
-phosphorylation destabilizes the glucose (give energy for reaction)
Product inhibition hexokinase
At high concentrations, G-6P inhibits hexokinase
How does glucose bind hexokinase
To the subtrate binding site and induced fit
Hexokinase isozymes
-Hexokinase 1: muscle and brain
> low Km: high affinity
> product inhibition by G-6P
-Hexokinase 4/ glucokinase
> high Km, low affinity
> not inhibited by G-6P
> In liver: glucose should not stay, only if there is excess glucose, it should be taken up and converted, because liver has to deal with it.
Where expression Hexokinase 1 and where glucokinase?
Glucokinase in pancreas, GI tract and liver: only use and retain glucose in the fed state
Hexokinase 1 in brain, adipose tissue and muscle tissue and erythrocytes: need glucose for energy and function.
Investment phase of glycolysis summed
Two phosphorylation steps
> Glucose to fructose-1,6-bisphosphate
-Glucose > G-6P (costs ATP, hexokinase)
-G-6P <=> Fructose 6-phosphate (phosphoglucose isomerase)
-F6-P > F-1,6-BP (costs ATP, phosphofructokinase)
Stage 2 of glycolysis
Fructose-1,6-bisphosphate (C6) is split into two C3 molecules by aldolase
- to DHAP (dyhydroxyacetone phosphate) and GAP (glyeraldehyde 3-phosphate)
Percentage products of conversion F-1,6-P by aldolase
96% DHAP
4% GAP > toxic, will attack proteins
Conversion glyceraldehyde 3-phosphate to dihydroxyacetone phosphate by the enzyme…
TIM enzyme: triose phosphate isomerase.
> diffusion controlled
> both sides reversible
> but GAP is the one used in stage 3
Stage 3 glycolysis
Oxidation steps to pyruvate
> yield 2 ATP and 1 NADH per C3 product
> 4 ATP produced in total
> net: 2 ATP production (2 invested)
> Take energy in electrons by transferring 1 hydride ion to NAD+ per C3 split product
Sum reaction glycolysis
Glucose + 2 Pi + 2 ADP + 2 NAD+ > 2 pyruvate + 2 ATP + 2 NADH + 2 H+ + 2 H2O
Fates pyruvate
-To acetyl-CoA, oxidation further in mitochondria
-To lacate to regenerate NAD+
-To ethanol via acetaldehyde (decarboxylation and reduction) to regenerate NAD+)
Anaerobic glycolysis
Making lactate from pyruvate using lactate dehydrogenase
Which steps in glycolysis are tightly regualted?
The irreversible ones
Regulated steps glycolysis: the reactions and enzymes
-Glucose> Glucose-6-P (hexokinase)
-Fructose -6-P > Fructose 1,6-bisphosphate (PFK-1)
-Phosphoenolpyruvate (PEP) > pyruvate (Pyruvate Kinase/ PK)
Thermodynamics at regulated steps glycolysis
Reactions with dG «_space;0
Regulation phosphofructokinase-1 in muscle
PFK-1
> promoted by high [AMP]
> inhibited by high [ATP], citrate, or low pH (anaerobic glycolysis)
-High ATP or citrate indicate high energy status of the cell
ATP/AMP ratio in cell through
Adenylate kinase
ADP + ADP <=> AMP + ATP
- high AMP: low energy status cell
-High ATP: high energy status cell
Regulation glycolysis in resting muscle
It is off!
High G-6-P inhibits HK
ATP and citrate inhibit PFK-1 and PK
Regulation glycolysis in active muscle
Glycolysis is on
> ATP/AMP ratio low (high AMP) promotes PFK-1
> F-1,6-BP promotes PK (feedforward stimulation)
Liver blood sugar homeostasis
If blood glucose is high, it still needs to take up glucose and hold it to make glycogen (no product inhibition by G-6-P)
> then inhibition gluconeogenesis and promotion glycogen storage
Regulation glycolysis in liver
-GK: low affinity for glucose, high KM. No product inhibition
-PFK-1: ATP regulation like in the muscle, no effect of low pH, but citrate does inhibit PFK-1 (enhances effect ATP)
Regulation PFK-1 in liver through PFK-2
PFK-2 can convert F-6P to F-2,6-BP.
-F-2,6-BP is a metabolite which regulates glycolysis/gluconeogensis balance in liver.
-F-2,6-BP produced by PFK2 and dephosphorylated by FBPase2
> 2 catalytic domains on one protein (PFK-2/FBPase2)
-F-2,6-BP activates PFK-1 (lower Km, slightly increases Vmax)
Regulation PK in liver
-Phosphorylated form is less active
-Dephosphorylated form in more active
-Phosphorylation PK in low blood glucose level and dephosphorylation in high blood glucose
-F-1,6-BP promotes PK
-ATP and alanine (indicate high energy status or low blood glucose level (starved state))
Other sugars which can enter glycolysis
-Fructose, galactose and lactose.
> Fructose in liver: to DHAP or GAP
> Fructose in adipose tissue: to F-6P
> Galactose to G-6P
Where is fructokinase expressed?
Only in the liver
> converts fructose to F-1P. (costs ATP)
> F-1P converted to glyceraldehyde and DHAP (F-1P aldolase)
> Glyceraldehyde ? GAP (triose kinase, costs ATP)
Why is fructose more fattening than glucose?
The most regulated step: PFK1 is bypassed in the liver
> glycolysis cannot be stopped
> TCA cycle will stop when enough energy, conversion to fat
Substrate pentose phosphate pathway (PPP)
Glucose-6-P
Product PPP
Ribose-5-P (C5)
Where does the PPP take place, and glycolysis
Both cytosol
Function PPP
Production NADPH and Ribose-5-P (for RNA/DNA synthesis)
Use of NADPH
Synthesis
-FA synthesis
-Cholesterol synthesis
-Neurotransmitter and nucleotide biosynthesis
Detoxification
-Reduction of oxidized glutathione (keep reduced environment, neutralize ROS)
-CYP enzymes
Two stages PPP
-Oxidative phase
-Non-oxidative phase
Oxidative phase PPP
-G6P > 6-PG (yield NADPH)
-6-PG > 6-PG lactone
-6-PG lactone > Ribulose-5-P (yield NADPH and CO2)
> sum: G6P + 2 NADP+ + H2O > R-5-P + 2 NADPH + 2 H+ + CO2
Regulation oxidative phase PPP
First step via NADP+ concentration
> G6P to 6-PG (glucose 6 phosphate dehydrogenase)
Non-oxidative phase PPP
Make C3,4,5,6,7 molecules from ribose-5-P (from ribulose-5-P, this conversion is oxidative)
Link PPP and glycolysis
the nonoxidative phase can yield glycolysis intermediates
C5 (ribose-5-P) + C5 <=> C3+C7 (by transketolase)
C3+C7 <=> C6+C4 (by transaldolase)
C4+C5 <=> C6 + C3
Sum:
3 C3 (ribose-5-P) <=> 2 C6 (fructose-6-P) + C3 (glyceraldehyde-3-P)
> intermediates glycolysis
Glucose fates in erythrocytes
NADPH is important to reduce oxidative stress
> 90% glucose to anaerobic glycolysis
> 10% to PPP
Function glutathione
Reduce oxidative stress in reduced form.
Reduced glutathione (GSH) reaction with ROOH (oxidative stress)
2 GSH (red) + ROOH > GSSG (condensed with disulfide bond, oxidized) + H2O + ROH
> Glutathione reductase reduces dimer GSSG back to monomers GDH using the electrons from NADPH from the PPP
ROS in red blood cells
Cause hemolysis
> when defect glucose-6-phosphate dehydrogenase (committed step PPP)
The PPP is flexible: explain
Requirements of the cell determine which product is produced (NADPH, ribose-5-phosphate, ATP)
HC06: Substrate TCA/Krebs cycle
Acetyl-CoA
Roads to acetyl-CoA
From: pyruvate, fatty acids, ketone bodies and some amino acids (and ethanol)
Ketone bodies derived from
acetyl-CoA
TCA cycle in the ..
mitochondrion
Anaerobic glycolysis uses LDH (lactate dehydrogenase) to generate
NAD+, to keep the glycolysis and ATP synthesis running
Each acetyl-CoA gives
3 NADH and 1 FADH2
NADH type and ATP yield
Soluble cofactor yields 2.5 ATP
FADH2 type and ATP yield
Prostethic group yields 1.5 ATP
How many CO2 are released per TCA cycle
2 CO2
> conversion isocitrate to a-ketoglutarate
> conversion a-ketoglutarate to succinyl-CoA
TCA cycle summary
Series redox reactions
> Oxidation acetyl group (C2) to two CO2 (acetyl-CoA is completely gone in the cycle and cannot be used for biosynthesis once in TCA cycle).
> harvest high energy electrons
> these are used for ATP synthesis
Thermodynamics TCA cycle
Condensation reaction oxaloacetate with acetyl-CoA to citrate
Substrate level phosphorylation of GDP to GTP in conversion succinyl-CoA to succinate
two oxidative decarboxylations
Which enzyme links glycolysis to TCA cycle. Why is this important?
The pyruvate dehydrogenase complex
> irreversible: regulated
> pyruvate to acetyl-CoA
Oxidative decarboxylation of pyruvate by PDH
Three subunit enzymes
E1: Pyruvate dehydrogenase
- Pyruvate > CO2 couped to TPP > Acyl-TPP
E2: Pyruvate transacetylase
-Acyl-TPP > TPP coupled to Lip-S-S > acyl-lipoate and acyl-lipoate> Lip-SH-SH coupled to CoASH to acetyl-CoA.
E3: dihydrolipoyl dehydrogenase: Lip-SH-SH (no disulfide bond)> Lip-S-S (disulfide bond) coupled to FAD > FADH2 and FADH2 > FAD coupled to NAD+ > NADH
What kind of molecule is TPP, the acyl carrier involved in the oxidative decarboxylation of pyruvate?
TPP: thiamine pyrophosphate (vitamin B1)
Regulation TCA cycle most important point
The PDH complex
> inactivation: phosphorylation by kinase
> activation; dephosphorylation by phosphatase
> Muscle: high ATP/ADP, Acetyl-CoA and high NADH inhibit PDH
> Muscle: low ATP/ADP (much ADP) and pyruvate promote PDH
Why is it important that ATP inhibits PDH
Conversion pyruvate to acetyl-CoA is irreversible and there is enough energy
> ability to still make glucose through gluconeogenesis
Intermediate formed in condensation reaction oxaloacetate (C4) + acetyl-CoA (C2) + H2O > citrate (C6) + CoA
Citryl-CoA
Adding TCA cycle intermediates
Anaplerosis
> more pyruvate used to keep the cycle going > more O2 uptake due to large amounts of NADH and FADH2 produced for oxidative phosphorylation
Succinyl-CoA can be used for biosynthesis of … and this is an example of …
Porphyrins and heme
> cataplerosis, removal TCA intermediate
Control points TCA cycle
Isocitrate dehydrogenase
> inhibited by ATP and NADH
> promoted by ADP
a-ketoglutarate dehydrogenase
> inhibited by ATP, succinyl-CoA and NADH
